Use of benzoxaboroles as volatile antimicrobial agents on meats, plants, or plant parts

ABSTRACT

This invention is related to use of a volatile antimicrobial compound against pathogens affecting meats, plants, or plant parts. The volatile antimicrobial compounds provided include certain oxaborole compounds, for example benzoxaboroles. Delivery systems are provided to take advantage of the volatile nature of these antimicrobial compounds. Also combinations with a volatile plant growth regulator, for example 1-methylcyclopropene, are disclosed.

CROSS-REFERENCE

This application is a continuation of U.S. patent application Ser. No.14/167,093, filed Jan. 29, 2014, and claims the benefit of U.S.provisional patent application No. 61/758,313 filed Jan. 30, 2013, eachof which is incorporated by reference herein in its entirety.

BACKGROUND OF THE INVENTION

A number of compounds containing an oxaborole ring have been disclosedpreviously. However, there has been no teaching that these oxaborolecompounds are volatile antimicrobial agents. In addition, theseoxaborole compounds have not been used in agricultural applications.

Thus, there remains a need to develop new uses of various volatileantimicrobial agents and/or combinations with a volatile plant growthregulator, in particular for agricultural applications.

SUMMARY OF THE INVENTION

This invention is related to the use of a volatile antimicrobialcompound against pathogens affecting meats, plants, or plant parts. Thevolatile antimicrobial compounds provided include certain oxaborolecompounds, for example benzoxaboroles. Delivery systems are provided totake advantage of the volatile nature of these antimicrobial compounds.Also combinations with a volatile plant growth regulator, for example1-methylcyclopropene (1-MCP), are disclosed.

In one aspect, provided is a method of using a volatile antimicrobialcompound against pathogens affecting meats, plants, or plant parts. Themethod comprises contacting the meats, plants, or plant parts with aneffective amount of the volatile antimicrobial compound having astructure of formula (I), (II), or (III):

wherein q1 and q2 are independently 1, 2, or 3;

q3=0, 1, 2, 3, or 4;

M is hydrogen, halogen, —OCH₃, or —CH₂—O—CH₂—O—CH₃;

M¹ is halogen, —CH₂OH, or —OCH₃;

X is O, S, or NR^(1c), wherein R^(1c) is hydrogen, substituted alkyl, orunsubstituted alkyl;

R¹, R^(1a), R^(1b), R², and R⁵ are independently hydrogen, OH, NH₂, SH,CN, NO₂, SO₂, OSO₂OH, OSO₂NH₂, substituted or unsubstituted cycloalkyl,substituted or unsubstituted heterocycloalkyl, substituted orunsubstituted aryl, or substituted or unsubstituted heteroaryl;

R* is substituted or unsubstituted aryl, substituted or unsubstitutedarylalkyl, substituted or unsubstituted heteroaryl, substituted orunsubstituted heteroarylalkyl, or substituted or unsubstituted vinyl;

with a proviso that when M is F, R* is not a member selected from:

and with a proviso that when M is Cl, R* is not a member selected from:

and with a proviso that when M is hydrogen, R* is not a member selectedfrom:

wherein s=1 or 2; and R³ and R⁴ are independently methyl or ethyl;

and with a provision that when M is OCH₃, R* is not a member selectedfrom:

and with a provision that when M¹ is F, R* is not a member selectedfrom:

and agriculturally acceptable salts thereof.

In one embodiment of the method provided, the pathogen is selected fromthe group consisting of Alternaria spp., Aspergillus spp., Botryospheriaspp., Botrytis spp., Byssochlamys spp., Colletotrichum spp., Diplodiaspp., Fusarium spp., Geotrichum spp., Lasiodiplodia spp., Monoliniaspp., Mucor spp., Penicillium spp., Pezicula spp., Phomopsis spp.,Phytophthora spp., Pythium spp., Rhizoctonia spp., Rhizopus spp.,Sclerotinia spp., and Venturia spp. In another embodiment, the pathogenis selected from the group consisting of Erwinia spp., Pectobacteriumspp., Pseudomonas spp., Ralstonia spp., Xanthomonas spp., Salmonellaspp., Escherichia spp., Listeria spp., Bacillus spp., Shigella spp., andStaphylococcus spp. In another embodiment, the pathogen is selected fromthe group consisting of Candida spp., Debaryomyces spp., Bacillus spp.,Campylobacter spp., Clostridium spp., Cryptosporidium spp., Giardiaspp., Vibrio spp., and Yersinia spp. In another embodiment, the methodcomprises a pre-harvest treatment or post-harvest treatment. In afurther embodiment, the pre-harvest treatment is selected from the groupconsisting of seed treatment and transplant treatment. In anotherembodiment, the post-harvest treatment is selected from the groupconsisting of treatment during field packing, treatment duringpalletization, in-box treatment, treatment during transportation, andtreatment during storage and/or throughout the distribution network.

In another embodiment, the plants or plant parts comprise transgenicplants or transgenic plant parts. In another embodiment, the plants orplant parts are selected from the group consisting of corn, wheat,cotton, rice, soybean, and canola. In another embodiment, the plants orplant parts are selected from the group consisting of fruit, vegetables,nursery, turf and ornamental crops. In a further embodiment, the fruitis selected from the group consisting of banana, pineapple, citrusincluding oranges, lemon, lime, grapefruit, and other citrus, grapes,watermelon, cantaloupe, muskmelon, and other melons, apple, peach, pear,cherry, kiwifruit, mango, nectarine, guava, papaya, persimmon,pomegranate, avocado, fig, and berries including strawberry, blueberry,raspberry, blackberry, currants and other types of berries. In a furtherembodiment, the vegetable is selected from the group consisting oftomato, potato, sweet potato, cassava, pepper, bell pepper, carrot,celery, squash, eggplant, cabbage, cauliflower, broccoli, asparagus,mushroom, onion, garlic, leek, and snap bean. A further embodiment, theflower or flower part is selected from the group consisting of roses,carnations, orchids, geraniums, lily or other ornamental flowers. Afurther embodiment, the meat is selected from the group of beef, bison,chicken, deer, goat, turkey, pork, sheep, fish, shellfish, mollusks, ordry-cured meat products.

In one embodiment, the contacting comprises applying the volatileantimicrobial compound by ways selected from the group consisting ofspray, mist, thermal or non-thermal fogging, drench, gas treatment, andcombinations thereof. In a further embodiment, the gas treatment isselected from the group consisting of release from a sachet, releasefrom a synthetic or natural film, fibrous material, and/or release fromliner or other packaging materials, release from powder, release from agas-releasing generator, release using a compressed or non-compressedgas cylinder, release from a droplet inside a box, and combinationsthereof. In another embodiment, the method further comprises contactingthe meats, plants, plant parts with a volatile plant growth regulator.In a further embodiment, the volatile plant growth regulator is acyclopropene compound. In a further embodiment, the cyclopropenecompound comprises 1-methylcyclopropene (1-MCP).

In another aspect, provided is a method of using a volatileantimicrobial compound against pathogens affecting meats, plants, orplant parts. The method comprises contacting the meats, plants, or plantparts with an effective amount of the volatile antimicrobial compound offormula (IV):

wherein A and D together with the carbon atoms to which they areattached form a 5-, 6-, or 7-membered fused ring which may besubstituted by C₁-C₆-alkyl, C₁-C₆-alkoxy, hydroxy, halogen, nitro,nitrile, amino, amino substituted by one or more C₁-C₆-alkyl groups,carboxy, acyl, aryloxy, carbonamido, carbonamido substituted byC₁-C₆-alkyl, sulfonamido or trifluoromethyl or the fused ring may linktwo oxaborole rings;

X is a group —CR⁷R⁸ wherein R⁷ and R⁸ are each independently hydrogen,C₁-C₆-alkyl, nitrile, nitro, aryl, arylalkyl or R⁷ and R⁸ together withthe carbon atom to which they are attached form an alicyclic ring; and

R⁶ is hydrogen, C₁-C₁₈-alkyl, (C₁-C₁₈-alkyl substituted by C₁-C₆-alkoxy,C₁-C₆-alkylthio, hydroxy, amino, amino substituted by C₁-C₁₈-alkyl,carboxy, aryl, aryloxy, carbonamido, carbonamido substituted byC₁-C₆-alkyl, aryl or arylalkyl, arylalkyl, aryl, heteroaryl, cycloalkyl,C₁-C₁₈-alkyleneamino, C₁-C₁₈-alkyleneamino substituted by phenyl,C₁-C₆-alkoxy or C₁-C₆-alkylthio, carbonyl alkyleneamino or a radical offormula (V):

wherein A, D and X are as defined herein except for boronophthalide;

and agriculturally acceptable salts thereof.

In one embodiment of the method provided, the pathogen is selected fromthe group consisting of Alternaria spp., Aspergillus spp., Botryospheriaspp., Botrytis spp., Byssochlamys spp., Colletotrichum spp., Diplodiaspp., Fusarium spp., Geotrichum spp., Lasiodiplodia spp., Monoliniaspp., Mucor spp., Penicillium spp., Pezicula spp., Phomopsis spp.,Phytophthora spp., Pythium spp., Rhizoctonia spp., Rhizopus spp.,Sclerotinia spp., and Venturia spp. In another embodiment, the pathogenis selected from the group consisting of Erwinia spp., Pectobacteriumspp., Pseudomonas spp., Ralstonia spp., Xanthomonas spp.; Salmonellaspp., Escherichia spp., Listeria spp., Bacillus spp., Shigella spp., andStaphylococcus spp. In another embodiment, the pathogen is selected fromthe group consisting of Candida spp., Debaryomyces spp., Bacillus spp.,Campylobacter spp., Clostridium spp., Cryptosporidium spp., Giardiaspp., Vibrio spp., and Yersinia spp. In another embodiment, the methodcomprises a pre-harvest treatment or post-harvest treatment. In afurther embodiment, the pre-harvest treatment is selected from the groupconsisting of seed treatment and transplant treatment. In anotherembodiment, the post-harvest treatment is selected from the groupconsisting of treatment during field packing, treatment duringpalletization, in-box treatment, treatment during transportation, andtreatment during storage and/or throughout the distribution network.

In another embodiment, the plants or plant parts comprise transgenicplants or transgenic plant parts. In another embodiment, the plants orplant parts are selected from the group consisting of corn, wheat,cotton, rice, soybean, and canola. In another embodiment, the plants orplant parts are selected from the group consisting of fruit, vegetables,nursery, turf and ornamental crops. In a further embodiment, the fruitis selected from the group consisting of banana, pineapple, citrusincluding oranges, lemon, lime, grapefruit, and other citrus, grapes,watermelon, cantaloupe, muskmelon, and other melons, apple, peach, pear,cherry, kiwifruit, mango, nectarine, guava, papaya, persimmon,pomegranate, avocado, fig, and berries including strawberry, blueberry,raspberry, blackberry, currants and other types of berries. In a furtherembodiment, the vegetable is selected from the group consisting oftomato, potato, sweet potato, cassava, pepper, bell pepper, carrot,celery, squash, eggplant, cabbage, cauliflower, broccoli, asparagus,mushroom, onion, garlic, leek, and snap bean. A further embodiment, theflower or flower part is selected from the group consisting of roses,carnations, orchids, geraniums, lily or other ornamental flowers. Afurther embodiment, the meat is selected from the group of beef, bison,chicken, deer, goat, turkey, pork, sheep, fish, shellfish, mollusks, ordry-cured meat products.

In one embodiment, the contacting comprises applying the volatileantimicrobial compound by ways selected from the group consisting ofspray, mist, thermal or non-thermal fogging, drench, gas treatment, andcombinations thereof. In a further embodiment, the gas treatment isselected from the group consisting of release from a sachet, releasefrom a synthetic or natural film, fibrous material, and/or release froma liner or other packaging materials, release from powder, release froma gas-releasing generator, release using a compressed or non-compressedgas cylinder, release from a droplet inside a box, and combinationsthereof. In another embodiment, the method further comprises contactingthe meats, plants, or plant parts with a volatile plant growthregulator. In a further embodiment, the volatile plant growth regulatoris a cyclopropene compound. In a further embodiment, the cyclopropenecompound comprises 1-methylcyclopropene (1-MCP).

In another aspect, provided is a method of using a volatileantimicrobial compound against pathogens affecting meats, plants, orplant parts. The method comprises contacting the meats, plants, or plantparts with an effective amount of the volatile antimicrobial compound offormula (VI):

wherein each R is independently hydrogen, alkyl, alkene, alkyne,haloalkyl, haloalkene, haloalkyne, alkoxy, alkeneoxy, haloalkoxy, aryl,heteroaryl, arylalkyl, arylalkene, arylalkyne, heteroarylalkyl,heteroarylalkene, heteroarylalkyne, halogen, hydroxyl, nitrile, amine,ester, carboxylic acid, ketone, alcohol, sulfide, sulfoxide, sulfone,sulfoximine, sulfilimine, sulfonamide, sulfate, sulfonate, nitroalkyl,amide, oxime, imine, hydroxylamine, hydrazine, hydrazone, carbamate,thiocarbamate, urea, thiourea, carbonate, aryloxy, or heteroaryloxy;

n=1, 2, 3, or 4;

B is boron;

X=(CR₂)_(m) where m=1, 2, 3, or 4;

Y is alkyl, alkene, alkyne, haloalkyl, haloalkene, haloalkyne, alkoxy,alkeneoxy, haloalkoxy, aryl, heteroaryl, arylalkyl, arylalkene,arylalkyne, heteroarylalkyl, heteroarylalkene, heteroarylalkyne,hydroxyl, nitrile, amine, ester, carboxylic acid, ketone, alcohol,sulfide, sulfoxide, sulfone, sulfoximine, sulfilimine, sulfonamide,sulfate, sulfonate, nitroalkyl, amide, oxime, imine, hydroxylamine,hydrazine, hydrazone, carbamate, thiocarbamate, urea, thiourea,carbonate, aryloxy, or heteroaryloxy;

with a proviso that R is not aryloxy or heteroaryloxy when Y ishydroxyl;

and agriculturally acceptable salts thereof.

In one embodiment, the volatile antimicrobial compound has a structureof formula (VII):

wherein W=(CH₂)_(q) where q is 1, 2, or 3.

In another embodiment, the volatile antimicrobial compound has astructure of

In one embodiment of the method provided, the pathogen is selected fromthe group consisting of Alternaria spp., Aspergillus spp., Botryospheriaspp., Botrytis spp., Byssochlamys spp., Colletotrichum spp., Diplodiaspp., Fusarium spp., Geotrichum spp., Lasiodiplodia spp., Monoliniaspp., Mucor spp., Penicillium spp., Pezicula spp., Phomopsis spp.,Phytophthora spp., Pythium spp., Rhizoctonia spp., Rhizopus spp.,Sclerotinia spp., and Venturia spp. In another embodiment, the pathogenis selected from the group consisting of Erwinia spp., Pectobacteriumspp., Pseudomonas spp., Ralstonia spp., Xanthomonas spp., Salmonellaspp., Escherichia spp., Listeria spp., Bacillus spp., Shigella spp., andStaphylococcus spp. In another embodiment, the pathogen is selected fromthe group consisting of Candida spp., Debaryomyces spp., Bacillus spp.,Campylobacter spp., Clostridium spp., Cryptosporidium spp., Giardiaspp., Vibrio spp., and Yersinia spp. In another embodiment, the methodcomprises a pre-harvest treatment or post-harvest treatment. In afurther embodiment, the pre-harvest treatment is selected from the groupconsisting of seed treatment and transplant treatment. In anotherembodiment, the post-harvest treatment is selected from the groupconsisting of treatment during field packing, treatment duringpalletization, in-box treatment, treatment during transportation, andtreatment during storage and/or throughout the distribution network.

In another embodiment, the plants or plant parts comprise transgenicplants or transgenic plant parts. In another embodiment, the plants orplant parts are selected from the group consisting of corn, wheat,cotton, rice, soybean, and canola. In another embodiment, the plants orplant parts are selected from the group consisting of fruit, vegetables,nursery, turf and ornamental crops. In a further embodiment, the fruitis selected from the group consisting of banana, pineapple, citrusincluding oranges, lemon, lime, grapefruit, and other citrus, grapes,watermelon, cantaloupe, muskmelon, and other melons, apple, peach, pear,cherry, kiwifruit, mango, nectarine, guava, papaya, persimmon,pomegranate, avocado, fig, and berries including strawberry, blueberry,raspberry, blackberry, currants and other types of berries. In a furtherembodiment, the vegetable is selected from the group consisting oftomato, potato, sweet potato, cassava, pepper, bell pepper, carrot,celery, squash, eggplant, cabbage, cauliflower, broccoli, asparagus,mushroom, onion, garlic, leek, and snap bean. A further embodiment, theflower or flower part is selected from the group consisting of roses,carnations, orchids, geraniums, lily or other ornamental flowers. Afurther embodiment, the meat is selected from the group of beef, bison,chicken, deer, goat, turkey, pork, sheep, fish, shellfish, mollusks, ordry-cured meat products.

In one embodiment, the contacting comprises applying the volatileantimicrobial compound by ways selected from the group consisting ofspray, mist, thermal or non-thermal fogging, drench, gas treatment, andcombinations thereof. In a further embodiment, the gas treatment isselected from the group consisting of release from a sachet, releasefrom a synthetic or natural film, fibrous material, and/or release fromliner or other packaging materials, release from powder, release from agas-releasing generator, release using a compressed or non-compressedgas cylinder, release from a droplet inside a box, and combinationsthereof. In another embodiment, the method further comprises contactingthe meats, plants, plant parts with a volatile plant growth regulator.In a further embodiment, the volatile plant growth regulator is acyclopropene compound. In a further embodiment, the cyclopropenecompound comprises 1-methylcyclopropene (1-MCP).

In another aspect, provided is a method of using a volatileantimicrobial compound against pathogens affecting meats, plants, orplant parts. The method comprises contacting the meats, plants, or plantparts with an effective amount of the volatile antimicrobial compound offormula (VIII):

wherein R^(a) is CN, C(O)NR⁹R¹⁰, or C(O)OR¹¹ wherein R¹¹ is hydrogen,substituted alkyl, or unsubstituted alkyl,

X is N, CH and CR^(b);

R^(b) is halogen, substituted or unsubstituted alkyl, C(O)R¹², C(O)OR¹²,OR¹², NR¹²R¹³, wherein R⁹, R¹⁰, R¹², and R¹³ are independently hydrogen,substituted or unsubstituted alkyl, substituted or unsubstitutedheteroalkyl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted heterocycloalkyl, substituted or unsubstituted aryl, orsubstituted or unsubstituted heteroaryl;

with a proviso that R⁹ and R¹⁰, together with the atoms to which theyare attached, are optionally combined to form a 4- to 8-memberedsubstituted or unsubstituted heterocycloalkyl ring;

and with a proviso that R¹² and R¹³, together with the atoms to whichthey are attached, are optionally combined to form a 4- to 8-memberedsubstituted or unsubstituted heterocycloalkyl ring;

and agriculturally acceptable salts thereof.

In one embodiment of the method provided, the pathogen is selected fromthe group consisting of Alternaria spp., Aspergillus spp., Botryospheriaspp., Botrytis spp., Byssochlamys spp., Colletotrichum spp., Diplodiaspp., Fusarium spp., Geotrichum spp., Lasiodiplodia spp., Monoliniaspp., Mucor spp., Penicillium spp., Pezicula spp., Phomopsis spp.,Phytophthora spp., Pythium spp., Rhizoctonia spp., Rhizopus spp.,Sclerotinia spp., and Venturia spp. In another embodiment, the pathogenis selected from the group consisting of Erwinia spp., Pectobacteriumspp., Pseudomonas spp., Ralstonia spp., Xanthomonas spp., Salmonellaspp., Escherichia spp., Listeria spp., Bacillus spp., Shigella spp., andStaphylococcus spp. In another embodiment, the pathogen is selected fromthe group consisting of Candida spp., Debaryomyces spp., Bacillus spp.,Campylobacter spp., Clostridium spp., Cryptosporidium spp., Giardiaspp., Vibrio spp., and Yersinia spp. In another embodiment, the methodcomprises a pre-harvest treatment or post-harvest treatment. In afurther embodiment, the pre-harvest treatment is selected from the groupconsisting of seed treatment and transplant treatment. In anotherembodiment, the post-harvest treatment is selected from the groupconsisting of treatment during field packing, treatment duringpalletization, in-box treatment, treatment during transportation, andtreatment during storage and/or throughout the distribution network.

In another embodiment, the plants or plant parts comprise transgenicplants or transgenic plant parts. In another embodiment, the plants orplant parts are selected from the group consisting of corn, wheat,cotton, rice, soybean, and canola. In another embodiment, the plants orplant parts are selected from the group consisting of fruit, vegetables,nursery, turf and ornamental crops. In a further embodiment, the fruitis selected from the group consisting of banana, pineapple, citrusincluding oranges, lemon, lime, grapefruit, and other citrus, grapes,watermelon, cantaloupe, muskmelon, and other melons, apple, peach, pear,cherry, kiwifruit, mango, nectarine, guava, papaya, persimmon,pomegranate, avocado, fig, and berries including strawberry, blueberry,raspberry, blackberry, currants and other types of berries. In a furtherembodiment, the vegetable is selected from the group consisting oftomato, potato, sweet potato, cassava, pepper, bell pepper, carrot,celery, squash, eggplant, cabbage, cauliflower, broccoli, asparagus,mushroom, onion, garlic, leek, and snap bean. A further embodiment, theflower or flower part is selected from the group consisting of roses,carnations, orchids, geraniums, lily or other ornamental flowers. Afurther embodiment, the meat is selected from the group of beef, bison,chicken, deer, goat, turkey, pork, sheep, fish, shellfish, mollusks, ordry-cured meat products.

In one embodiment, the contacting comprises applying the volatileantimicrobial compound by ways selected from the group consisting ofspray, mist, thermal or non-thermal fogging, drench, gas treatment, andcombinations thereof. In a further embodiment, the gas treatment isselected from the group consisting of release from a sachet, releasefrom a synthetic or natural film, release from liner or other packagingmaterials, release from powder, release from a gas-releasing generator,release using a compressed or non-compressed gas cylinder, release froma droplet inside a box, and combinations thereof. In another embodiment,the method further comprises contacting the meats, plants, plant partswith a volatile plant growth regulator. In a further embodiment, thevolatile plant growth regulator is a cyclopropene compound. In a furtherembodiment, the cyclopropene compound comprises 1-methylcyclopropene(1-MCP).

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the chemical structure of an exemplary Compound A of theinvention.

FIG. 2 shows the chemical structure of an exemplary Compound B of theinvention.

FIG. 3 shows fourteen compounds tested in Example 2.

FIGS. 4A-4D show representative photos of exemplary in vivo inhibitionresults in a green table grape inoculated with Botrytis cinerea andtreated using Compound A as described in Example 4, where 0.04 mg ofCompound A shows 100% inhibition and 0.0024 mg of Compound A shows noinhibition. FIG. 4A shows a grape sample not inoculated with Botrytiscinerea.

FIG. 4B shows an inoculated grape sample treated with 0.04 mg ofCompound A after the 14 day bioassay test described in Example 4.

FIG. 4C shows an inoculated grape sample treated with 0.0024 mg ofCompound A after the 14 day bioassay test.

FIG. 4D shows an inoculated grape sample treated with pure acetone,including no Compound A, after the 14 day bioassay test.

FIGS. 5A and 5B show representative photos of exemplary in vivoinhibition results in a strawberry inoculated with Botrytis cinerea andtreated using Compound A as described in Example 6, where 0.125 mg ofCompound A shows 100% inhibition. FIG. 5A shows a representative photoof exemplary in vivo inhibition of Botrytis cinerea by a volatileapplication of Compound A after a 3-day treatment at 21° C., followed byan additional 2 days at 21° C., as described in Example 6.

FIG. 5B shows an inoculated strawberry sample treated with pure acetone,including no Compound A, after the 3-day treatment at 21° C., followedby an additional 2 days at 21° C.

DETAILED DESCRIPTION OF THE INVENTION

Unless otherwise stated, the following terms used in this application,including the specification and claims, have the definitions givenbelow. It must be noted that, as used in the specification and theappended claims, the singular forms “a,” “an” and “the” include pluralreferents unless the context clearly dictates otherwise. Definition ofstandard chemistry terms may be found in reference works, includingCarey and Sundberg, Advanced Organic Chemistry 4^(th) Ed., Vols. A(2000) and B (2001), Plenum Press, New York, N.Y.

As used herein, the phrase “moiety” refers to a specific segment orfunctional group of a molecule. Chemical moieties are often recognizedchemical entities embedded in or appended to a molecule.

As used herein, the phrases “heteroatom” and “hetero-” refer to atomsother than carbon (C) and hydrogen (H). Examples of heteroatoms includeoxygen (O), nitrogen (N) sulfur (S), silicon (Si), germanium (Ge),aluminum (Al) and boron (B).

As used herein, the phrases “halo” and “halogen” are interchangeable andrefer to fluoro (—F), chloro (—Cl), bromo (—Br), and iodo (—I).

As used herein, the phrase “alkyl” refers to an unsubstituted orsubstituted, hydrocarbon group and can include straight, branched,cyclic, saturated and/or unsaturated features. Although the alkyl moietymay be an “unsaturated alkyl” moiety, which means that it contains atleast one alkene or alkyne moiety, typically, the alkyl moiety is a“saturated alkyl” group, which means that it does not contain any alkeneor alkyne moieties. Likewise, although the alkyl moiety may be cyclic,the alkyl moiety typically is acyclic group. Thus, in some embodiments,“alkyl” refers to an optionally substituted straight-chain, oroptionally substituted branched-chain saturated hydrocarbon monoradicalhaving from about one to about thirty carbon atoms in some embodiments,from about one to about fifteen carbon atoms in some embodiments, andfrom about one to about six carbon atoms in further embodiments.Examples of saturated alkyl radicals include, but are not limited to,methyl, ethyl, n-propyl, isopropyl, 2-methyl-1-propyl,2-methyl-2-propyl, 2-methyl-1-butyl, 3-methyl-1-butyl, 2-methyl-3-butyl,2,2-dimethyl-1-propyl, 2-methyl-1-pentyl, 3-methyl-1-pentyl,4-methyl-1-pentyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl,4-methyl-2-pentyl, 2,2-dimethyl-1-butyl, 3,3-dimethyl-1-butyl,2-ethyl-1-butyl, butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl,isopentyl, neopentyl, and n-hexyl, and longer alkyl groups, such asheptyl, and octyl. It should be noted that whenever it appears herein, anumerical range such as “1 to 6” refers to each integer in the givenrange; e.g., “1 to 6 carbon atoms” or “C₁₋₆” or “C₁-C₆” means that thealkyl group may consist of 1 carbon atom, 2 carbon atoms, 3 carbonatoms, 4 carbon atoms, 5 carbon atoms, and/or 6 carbon atoms, althoughthe present definition also covers the occurrence of the term “alkyl”where no numerical range is designated.

As used herein, the phrase “substituted alkyl” refers to an alkyl group,as defined herein, in which one or more (up to about five, preferably upto about three) hydrogen atoms is replaced by a substituentindependently selected from the substituent group defined herein.

As used herein, the phrases “substituents” and “substituted” refer togroups which may be used to replace another group on a molecule. Suchgroups are known to those of skill in the chemical arts and may include,without limitation, one or more of the following independently selectedgroups, or designated subsets thereof: halogen, —CN, —OH, —NO₂, —N₃, ═O,═S, ═NH, —SO₂, —NH₂, —COOH, nitroalkyl, amino, including mono- anddi-substituted amino groups, cyanato, isocyanato, thiocyanato,isothiocyanato, guanidinyl, O-carbamyl, N-carbamyl, thiocarbamyl, uryl,isouryl, thiouryl, isothiouryl, mercapto, sulfanyl, sulfinyl, sulfonyl,sulfonamidyl, phosphonyl, phosphatidyl, phosphoramidyl, dialkylamino,diarylamino, diarylalkylamino; and the protected compounds thereof. Theprotecting groups that may form the protected compounds of the abovesubstituents are known to those of skill in the art and may be found inreferences such as Greene and Wuts, Protective Groups in OrganicSynthesis, 3rd ed.; John Wiley & Sons, New York, N.Y. (1999) andKocienski, Protective Groups; Thieme Verlag, New York, N.Y. (1994) whichare incorporated herein by reference in their entirety.

As used herein, the phrase “alkoxy” refers to the group —O-alkyl, wherealkyl is as defined herein. In one embodiment, alkoxy groups include,e.g., methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy,sec-butoxy, n-pentoxy, n-hexoxy, 1,2-dimethylbutoxy, and the like. Thealkoxy can be unsubstituted or substituted.

As used herein, the phrases “cyclic” and “membered ring” refer to anycyclic structure, including alicyclic, heterocyclic, aromatic,heteroaromatic and polycyclic fused or non-fused ring systems asdescribed herein. The term “membered” is meant to denote the number ofskeletal atoms that constitute the ring. Thus, for example, pyridine,pyran, and pyrimidine are six-membered rings and pyrrole,tetrahydrofuran, and thiophene are five-membered rings.

As used herein, the phrase “aromatic” refers to a cyclic or polycyclicmoiety having a conjugated unsaturated (4n+2)π electron system (where nis a positive integer), sometimes referred to as a delocalized πelectron system.

As used herein, the phrase “aryl” refers to an optionally substituted,aromatic, cyclic, hydrocarbon monoradical of from six to about twentyring atoms, preferably from six to about ten carbon atoms and includesfused (or condensed) and non-fused aromatic rings. A fused aromatic ringradical contains from two to four fused rings where the ring ofattachment is an aromatic ring, and the other individual rings withinthe fused ring may be cycloalkyl, cycloalkenyl, cycloalkynyl,heterocycloalkyl, heterocycloalkenyl, heterocycloalkynyl, aromatic,heteroaromatic or any combination thereof. A non-limiting example of asingle ring aryl group includes phenyl; a fused ring aryl group includesnaphthyl, anthryl, azulenyl; and a non-fused bi-aryl group includesbiphenyl.

As used herein, the phrase “substituted aryl” refers to an aryl group,as defined herein, in which one or more (up to about five, preferably upto about three) hydrogen atoms is replaced by a substituentindependently selected from the group defined herein, (except asotherwise constrained by the definition for the aryl substituent).

As used herein, the phrase “heteroaryl” refers to an optionallysubstituted, aromatic, cyclic monoradical containing from about five toabout twenty skeletal ring atoms, preferably from five to about ten ringatoms and includes fused (or condensed) and non-fused aromatic rings,and which have one or more (one to ten, preferably about one to aboutfour) ring atoms selected from an atom other than carbon (i.e., aheteroatom) such as, for example, oxygen, nitrogen, sulfur, selenium,phosphorus or combinations thereof. The term heteroaryl includesoptionally substituted fused and non-fused heteroaryl radicals having atleast one heteroatom. A fused heteroaryl radical may contain from two tofour fused rings where the ring of attachment is a heteroaromatic ringand the other individual rings within the fused ring system may bealicyclic, heterocyclic, aromatic, heteroaromatic or any combinationthereof. The term heteroaryl also includes fused and non-fusedheteroaryls having from five to about twelve skeletal ring atoms, aswell as those having from five to about ten skeletal ring atoms.Examples of heteroaryl groups include, but are not limited to,acridinyl, benzo[1,3]dioxole, benzimidazolyl, benzindazolyl,benzoisooxazolyl, benzokisazolyl, benzofuranyl, benzofurazanyl,benzopyranyl, benzothiadiazolyl, benzothiazolyl, benzo[b]thienyl,benzothiophenyl, benzothiopyranyl, benzotriazolyl, benzoxazolyl,carbazolyl, carbolinyl, chromenyl, cinnolinyl, furanyl, furazanyl,furopyridinyl, furyl, imidazolyl, indazolyl, indolyl, indolidinyl,indolizinyl, isobenzofuranyl, isoindolyl, isoxazolyl, isoquinolinyl,isothiazolyl, naphthylidinyl, naphthyridinyl, oxadiazolyl, oxazolyl,phenoxazinyl, phenothiazinyl, phenazinyl, phenoxathiynyl, thianthrenyl,phenathridinyl, phenathrolinyl, phthalazinyl, pteridinyl, purinyl,puteridinyl, pyrazyl, pyrazolyl, pyridyl, pyridinyl, pyridazinyl,pyrazinyl, pyrimidinyl, pyrimidyl, pyrrolyl, quinazolinyl, quinolinyl,quinoxalinyl, tetrazolyl, thiadiazolyl, thiazolyl, thienyl, triazinyl,(1,2,3,)- and (1,2,4)-triazolyl and the like, and their oxides whereappropriate, such as for example pyridyl-N-oxide.

As used herein, the phrase “substituted heteroaryl” refers to aheteroaryl group, as defined herein, in which one or more (up to aboutfive, preferably up to about three) hydrogen atoms is replaced by asubstituent independently selected from the group defined herein.

As used herein, the phrase “leaving group” refers to a group with themeaning conventionally associated with it in synthetic organicchemistry, i.e., an atom or group displaceable under substitutionreaction conditions. Examples of leaving groups include, but are notlimited to, halogen, alkane- or arylenesulfonyloxy, such asmethanesulfonyloxy, ethanesulfonyloxy, thiomethyl, benzenesulfonyloxy,tosyloxy, and thienyloxy, dihalophosphinoyloxy, optionally substitutedbenzyloxy, isopropyloxy, acyloxy, and the like. In some embodiments, aleaving group can be HC(O)—COOH or RC(O)—COOH, wherein R is a C₁-C₆alkyl or substituted C₁-C₆ alkyl.

The compounds of the invention as described herein may be synthesizedusing standard synthetic techniques known to those of skill in the artor using methods known in the art in combination with methods describedherein. The starting materials used for the synthesis of the compoundsof the invention as described herein, can be obtained from commercialsources, such as Aldrich Chemical Co. (Milwaukee, Wis.), Sigma ChemicalCo. (St. Louis, Mo.), or the starting materials can be synthesized. Thecompounds described herein, and other related compounds having differentsubstituents can be synthesized using techniques and materials known tothose of skill in the art, such as described, for example, in March,Advanced Organic Chemistry 4^(th) Ed. (1992) John Wiley & Sons, NewYork, N.Y.; Carey and Sundberg, Advanced Organic Chemistry 4^(th) Ed.,Vols. A (2000) and B (2001) Plenum Press, New York, N.Y. and Greene andWuts, Protective Groups in Organic Synthesis, 3rd Ed. (1999) John Wiley& Sons, New York, N.Y., (all of which are incorporated by reference intheir entirety). General methods for the preparation of compounds asdisclosed herein may be derived from known reactions in the field, andthe reactions may be modified by the use of appropriate reagents andconditions, as would be recognized by the skilled person, for theintroduction of the various moieties found in the formulae as providedherein. For example, the compounds described herein can be modifiedusing various electrophiles or nucleophiles to form new functionalgroups or substituents.

In some embodiments, the volatile antimicrobial compound of theinvention has a structure of formula (I), (II), or (III):

wherein q1 and q2 are independently 1, 2, or 3;

q3=0, 1, 2, 3, or 4;

M is hydrogen, halogen, —OCH₃, or —CH₂—O—CH₂—O—CH₃;

M¹ is halogen, —CH₂OH, or —OCH₃;

X is O, S, or NR^(1c), wherein R^(1c) is hydrogen, substituted alkyl, orunsubstituted alkyl;

R¹, R^(1a), R^(1b), R², and R⁵ are independently hydrogen, OH, NH₂, SH,CN, NO₂, SO₂, OSO₂OH, OSO₂NH₂, substituted or unsubstituted cycloalkyl,substituted or unsubstituted heterocycloalkyl, substituted orunsubstituted aryl, or substituted or unsubstituted heteroaryl;

R* is substituted or unsubstituted aryl, substituted or unsubstitutedarylalkyl, substituted or unsubstituted heteroaryl, substituted orunsubstituted heteroarylalkyl, or substituted or unsubstituted vinyl;

with a proviso that when M is F, R* is not a member selected from:

and with a proviso that when M is Cl, R* is not a member selected from:

and with a proviso that when M is hydrogen, R* is not a member selectedfrom:

wherein s=1 or 2; and R³ and R⁴ are independently methyl or ethyl;

and with a provision that when M is OCH₃, R* is not a member selectedfrom:

and with a provision that when M¹ is F, R* is not a member selectedfrom:

and agriculturally acceptable salts thereof.

In one embodiment, the R* has a structure selected from:

wherein X is a member selected from CH═CH, N═CH, NR¹⁴, O and S;

wherein R¹⁴ is a member selected from H, substituted or unsubstitutedalkyl, substituted or unsubstituted aryl and substituted orunsubstituted arylalkyl;

Y is a member selected from CH and N;

R¹⁷ and R¹⁸ are members independently selected from H, substituted orunsubstituted alkyl, substituted or unsubstituted aryl, substituted orunsubstituted arylalkyl, (CH₂)_(V)OH, (CH₂)_(W)NR¹⁵R¹⁶, CO₂H, CO₂-alkyl,CONH₂, S-alkyl, S-aryl, SO-alkyl, SO-aryl, SO₂-alkyl, SO₂-aryl, SO₂H,SCF₂, CN, halogen, CF₃ and NO₂;

wherein R¹⁵ and R¹⁶ are members independently selected from hydrogen,substituted or unsubstituted alkyl and substituted or unsubstitutedalkanoyl;

v=1, 2, or 3; and

w=0, 1, 2, or 3.

In another embodiment, the R* has the following structure:

wherein R¹⁷, R¹⁸, R¹⁹, R²⁰, and R²¹ are independently selected from H,substituted or unsubstituted alkyl, substituted or unsubstituted aryl,substituted or unsubstituted arylalkyl, substituted or unsubstitutedalkyloxy, substituted or unsubstituted aryloxy, substituted orunsubstituted oxazolidin-2-yl, (CH₂)_(t)OH, CO₂H, CO₂-alkyl, CONH₂,CONH-alkyl, CON(alkyl)₂, OH, SH, S-alkyl, S-aryl, SO-alkyl, SO-aryl,SO₂-alkyl, SO₂-aryl, SO₂H, SCF₃, CN, halogen, CF₃, NO₂, (CH₂)—NR²²R²³,SO₂NH₂, OCH₂CH₂NH₂, OCH₂CH₂NH-alkyl and OCH₂CH₂N(alkyl)₂;

wherein t=1, 2 or 3;

u=0, 1, or 2;

R²² and R²³ are independently selected from H, substituted orunsubstituted alkyl, and substituted or unsubstituted alkanoyl.

In another embodiment, the R* has the following structure:

wherein R¹⁷, R¹⁸, R¹⁹, R²⁰, and R²¹ are independently selected from H,substituted or unsubstituted alkyl, substituted or unsubstituted aryl,substituted or unsubstituted arylalkyl, substituted or unsubstitutedalkyloxy, substituted or unsubstituted aryloxy, substituted orunsubstituted oxazolidin-2-yl, (CH₂)_(t)OH, CO₂H, CO₂-alkyl, CONH₂,CONH-alkyl, CON(alkyl)₂, OH, SH, S-alkyl, S-aryl, SO-alkyl, SO-aryl,SO₂-alkyl, SO₂-aryl, SO₂H, SCF₃, CN, halogen, CF₃, NO₂,(CH₂)_(u)NR²²R²³, SO₂NH₂, OCH₂CH₂NH₂, OCH₂CH₂NH-alkyl andOCH₂CH₂N(alkyl)₂;

wherein t=1, 2 or 3;

u=0, 1, or 2;

R²² and R²³ are independently selected from H, substituted orunsubstituted alkyl, and substituted or unsubstituted alkanoyl;

R²⁴ and R²⁵ are independently selected from H, substituted orunsubstituted alkyl, substituted or unsubstituted aryl, substituted orunsubstituted arylalkyl, substituted or unsubstituted alkyloxy,substituted or unsubstituted aryloxy, substituted or unsubstitutedoxazolidin-2-yl, (CH₂), OH, CO₂H, CO₂-alkyl, CONH₂, CONH-alkyl,CON(alkyl)₂, OH, SH, S-alkyl, S-aryl, SO-alkyl, SO-aryl, SO₂-alkyl,SO₂-aryl, SO₃H, SCF₃, CN, halogen, CF₃, NO₂, (CH₂)_(u)NR²²R²³, SO₂NH₂,OCH₂CH₂NH₂, OCH₂CH₂NH-alkyl and OCH₂CH₂N(alkyl)₂;

Z=1, 2, 3, 4, 5, or 6.

Additional antimicrobial compounds are also disclosed previously in U.S.Pat. No. 8,106,031, and International Patent Application WO2007/131072A2, the contents of which are hereby incorporated byreference in their entireties.

In some embodiments, the volatile antimicrobial compound of theinvention has the structure of formula (IV):

wherein A and D together with the carbon atoms to which they areattached form a 5-, 6-, or 7-membered fused ring which may besubstituted by C₁-C₆-alkyl, C₁-C₆-alkoxy, hydroxy, halogen, nitro,nitrile, amino, amino substituted by one or more C₁-C₆-alkyl groups,carboxy, acyl, aryloxy, carbonamido, carbonamido substituted byC₁-C₆-alkyl, sulfonamido or trifluoromethyl or the fused ring may linktwo oxaborole rings;

X is a group —CR⁷R⁸ wherein R⁷ and R⁸ are each independently hydrogen,C₁-C₆-alkyl, nitrile, nitro, aryl, arylalkyl or R⁷ and R⁸ together withthe carbon atom to which they are attached form an alicyclic ring; and

R⁶ is hydrogen, C₁-C₁₈-alkyl, C₁-C₁₈-alkyl substituted by C₁-C₆-alkoxy,C₁-C₆-alkylthio, hydroxy, amino, amino substituted by C₁-C₁₈-alkyl,carboxy, aryl, aryloxy, carbonamido, carbonamido substituted byC₁-C₆-alkyl, aryl or arylalkyl, arylalkyl, aryl, heteroaryl, cycloalkyl,C₁-C₁₈-alkyleneamino, C₁-C₁₈-alkyleneamino substituted by phenyl,C₁-C₆-alkoxy or C₁-C₆-alkylthio, carbonyl alkyleneamino or a radical offormula (V):

wherein A, D and X are as defined herein before except forboronophthalide;

and agriculturally acceptable salts thereof.

In one embodiment, the volatile antimicrobial compound of the inventionhas the structure of formula (IX):

wherein A, D, and X are defined as above;

Y is a divalent alkylene linking group containing up to 18 carbon atomsor a divalent alkylene linking group containing up to 18 carbon atomswhich is substituted by phenyl, C₁-C₆ alkoxy, C₁-C₆-alkylthio; carbonylalkylene amino; and

R³ and R⁴ are each, independently, hydrogen, C₁-C₁₈-alkyl or phenyl orR³ together with Y or part of Y forms a 5-, 6- or 7-membered ringcontaining the nitrogen atom.

In another embodiment, the volatile antimicrobial compound of theinvention has the structure of formula (X):

wherein A, D, and X are defined as above;

n is 1, 2, or 3;

R³ is hydrogen, C₁-C₁₈-alkyl or phenyl; and

R⁵ and R⁶ are each, independently, hydrogen, alkyl containing up to atotal of 16 carbon atoms or phenyl.

Additional antimicrobial compounds are also disclosed previously in U.S.Pat. No. 5,880,188, the content of which is hereby incorporated byreference in its entirety.

In another aspect, provided is a method of using a volatileantimicrobial compound against pathogens affecting meats, plants, orplant parts. The method comprises contacting the meats, plants, or plantparts with an effective amount of the volatile antimicrobial compound offormula (VI):

wherein each R is independently hydrogen, alkyl, alkene, alkyne,haloalkyl, haloalkene, haloalkyne, alkoxy, alkeneoxy, haloalkoxy, aryl,heteroaryl, arylalkyl, arylalkene, arylalkyne, heteroarylalkyl,heteroarylalkene, heteroarylalkyne, halogen, hydroxyl, nitrile, amine,ester, carboxylic acid, ketone, alcohol, sulfide, sulfoxide, sulfone,sulfoximine, sulfilimine, sulfonamide, sulfate, sulfonate, nitroalkyl,amide, oxime, imine, hydroxylamine, hydrazine, hydrazone, carbamate,thiocarbamate, urea, thiourea, carbonate, aryloxy, or heteroaryloxy;

n=1, 2, 3, or 4;

B is boron;

X=(CR₂)_(m) where m=1, 2, 3, or 4;

Y is alkyl, alkene, alkyne, haloalkyl, haloalkene, haloalkyne, alkoxy,alkeneoxy, haloalkoxy, aryl, heteroaryl, arylalkyl, arylalkene,arylalkyne, heteroarylalkyl, heteroarylalkene, heteroarylalkyne,hydroxyl, nitrile, amine, ester, carboxylic acid, ketone, alcohol,sulfide, sulfoxide, sulfone, sulfoximine, sulfilimine, sulfonamide,sulfate, sulfonate, nitroalkyl, amide, oxime, imine, hydroxylamine,hydrazine, hydrazone, carbamate, thiocarbamate, urea, thiourea,carbonate, aryloxy, or heteroaryloxy;

with a proviso that R is not aryloxy or heteroaryloxy when Y ishydroxyl;

and agriculturally acceptable salts thereof.

In one embodiment, the volatile antimicrobial compound has a structureof formula (VII):

wherein W=(CH₂)_(q) where q is 1, 2, or 3.

In another embodiment, the volatile antimicrobial compound has astructure of

In one embodiment of the method provided, the pathogen is selected fromthe group consisting of Alternaria spp., Aspergillus spp., Botryospheriaspp., Botrytis spp., Byssochlamys spp., Colletotrichum spp., Diplodiaspp., Fusarium spp., Geotrichum spp., Lasiodiplodia spp., Monoliniaspp., Mucor spp., Penicillium spp., Pezicula spp., Phomopsis spp.,Phytophthora spp., Pythium spp., Rhizoctonia spp., Rhizopus spp.,Sclerotinia spp., and Venturia spp. In another embodiment, the pathogenis selected from the group consisting of Erwinia spp., Pectobacteriumspp., Pseudomonas spp., Ralstonia spp., Xanthomonas spp.; Salmonellaspp., Escherichia spp., Listeria spp., Bacillus spp., Shigella spp., andStaphylococcus spp. In another embodiment, the pathogen is selected fromthe group consisting of Candida spp., Debaryomyces spp., Bacillus spp.,Campylobacter spp., Clostridium spp., Cryptosporidium spp., Giardiaspp., Vibrio spp., and Yersinia spp. In another embodiment, the methodcomprises a pre-harvest treatment or post-harvest treatment. In afurther embodiment, the pre-harvest treatment is selected from the groupconsisting of seed treatment and transplant treatment. In anotherembodiment, the post-harvest treatment is selected from the groupconsisting of treatment during field packing, treatment duringpalletization, in-box treatment, treatment during transportation, andtreatment during storage and/or throughout the distribution network.

In another embodiment, the plants or plant parts comprise transgenicplants or transgenic plant parts. In another embodiment, the plants orplant parts are selected from the group consisting of corn, wheat,cotton, rice, soybean, and canola. In another embodiment, the plants orplant parts are selected from the group consisting of fruit, vegetables,nursery, turf and ornamental crops. In a further embodiment, the fruitis selected from the group consisting of banana, pineapple, citrusincluding oranges, lemon, lime, grapefruit, and other citrus, grapes,watermelon, cantaloupe, muskmelon, and other melons, apple, peach, pear,cherry, kiwifruit, mango, nectarine, guava, papaya, persimmon,pomegranate, avocado, fig, and berries including strawberry, blueberry,raspberry, blackberry, currants and other types of berries. In a furtherembodiment, the vegetable is selected from the group consisting oftomato, potato, sweet potato, cassava, pepper, bell pepper, carrot,celery, squash, eggplant, cabbage, cauliflower, broccoli, asparagus,mushroom, onion, garlic, leek, and snap bean. A further embodiment, theflower or flower part is selected from the group consisting of roses,carnations, orchids, geraniums, lily or other ornamental flowers. Afurther embodiment, the meat is selected from the group of beef, bison,chicken, deer, goat, turkey, pork, sheep, fish, shellfish, mollusks, ordry-cured meat products.

In one embodiment, the contacting comprises applying the volatileantimicrobial compound by ways selected from the group consisting ofspray, mist, thermal or non-thermal fogging, drench, gas treatment, andcombinations thereof. In a further embodiment, the gas treatment isselected from the group consisting of release from a sachet, releasefrom a synthetic or natural film, fibrous material, and/or release froma liner or other packaging materials, release from powder, release froma gas-releasing generator, release using a compressed or non-compressedgas cylinder, release from a droplet inside a box, and combinationsthereof. In another embodiment, the method further comprises contactingthe meats, plants, plant parts with a cyclopropene compound. In afurther embodiment, the cyclopropene compound comprises1-methylcyclopropene (1-MCP).

In another aspect, provided is a method of using a volatileantimicrobial compound against pathogens affecting meats, plants, orplant parts. The method comprises contacting the meats, plants, or plantparts with an effective amount of the volatile antimicrobial compound offormula (VIII):

wherein R^(a) is CN, C(O)NR⁹R¹⁰, or C(O)OR¹¹ wherein R¹¹ is hydrogen,substituted alkyl, or unsubstituted alkyl,

X is N, CH and CR^(b);

R^(b) is halogen, substituted or unsubstituted alkyl, C(O)R¹², C(O)OR¹²,OR¹², NR¹²R¹³, wherein R⁹, R¹⁰, R¹², and R¹³ are independently hydrogen,substituted or unsubstituted alkyl, substituted or unsubstitutedheteroalkyl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted heterocycloalkyl, substituted or unsubstituted aryl, orsubstituted or unsubstituted heteroaryl;

with a proviso that R⁹ and R¹⁰, together with the atoms to which theyare attached, are optionally combined to form a 4- to 8-memberedsubstituted or unsubstituted heterocycloalkyl ring;

and with a proviso that R¹² and R¹³, together with the atoms to whichthey are attached, are optionally combined to form a 4- to 8-memberedsubstituted or unsubstituted heterocycloalkyl ring;

and agriculturally acceptable salts thereof.

In one embodiment, the volatile antimicrobial compound of the inventionhas the structure of formula (XI):

In another embodiment, the volatile antimicrobial compound of theinvention is selected from:

In another embodiment, the volatile antimicrobial compound of theinvention is selected from:

In another embodiment, the volatile antimicrobial compound of theinvention is selected from:

In one embodiment, the volatile antimicrobial compound of the inventionhas the structure of formula (XII):

In another embodiment, the volatile antimicrobial compound of theinvention is selected from:

wherein R³ is hydrogen, substituted or unsubstituted alkyl, orsubstituted or unsubstituted heteroalkyl.

In another embodiment, the volatile antimicrobial compound of theinvention is selected from:

wherein R³ is hydrogen, substituted or unsubstituted alkyl, orsubstituted or unsubstituted heteroalkyl.

In another embodiment, the volatile antimicrobial compound of theinvention is selected from:

wherein R³ is hydrogen, substituted or unsubstituted alkyl, orsubstituted or unsubstituted heteroalkyl.

In one embodiment, the volatile antimicrobial compound of the inventionhas the structure of formula (XIII):

wherein each of R¹ and R² is independently hydrogen, substituted orunsubstituted alkyl, or substituted or unsubstituted heteroalkyl.

In another embodiment, the volatile antimicrobial compound of theinvention is selected from:

In another embodiment, the volatile antimicrobial compound of theinvention is selected from:

wherein each of R¹ and R² is independently hydrogen, substituted orunsubstituted alkyl, or substituted or unsubstituted heteroalkyl.

In another embodiment, the volatile antimicrobial compound of theinvention is selected from:

wherein each of R¹ and R² is independently hydrogen, substituted orunsubstituted alkyl, or substituted or unsubstituted heteroalkyl.

In one embodiment, R^(b) is selected from fluorine and chlorine. Inanother embodiment, R^(b) is selected from OR²⁶ and NR²⁷R²⁸. In anotherembodiment when R^(b) is OR²⁶, R²⁶ is selected from H, substituted orunsubstituted alkyl, substituted or unsubstituted heteroalkyl,substituted or unsubstituted cycloalkyl, substituted or unsubstitutedheterocycloalkyl, substituted or unsubstituted aryl, and substituted orunsubstituted heteroaryl. In another embodiment when R^(b) is OR²⁶, R²⁶is selected from H, substituted or unsubstituted alkyl, substituted orunsubstituted heteroalkyl and substituted or unsubstituted cycloalkyl.In another embodiment when R^(b) is OR²⁶, R²⁶ is unsubstituted C₁-C₆alkyl. In another embodiment when R^(b) is OR²⁶, R²⁶ is unsubstitutedcycloalkyl. In another embodiment when R^(b) is OR²⁶, R²⁶ is alkyl,substituted with a member selected from substituted or unsubstitutedC₁-C₆ alkoxy. In another embodiment when R^(b) is OR²⁶, R²⁶ is alkyl,substituted with at least one halogen. In another embodiment when R^(b)is OR²⁶, R²⁶ is alkyl, substituted with at least one oxo moiety.

In another embodiment when R^(b) is OR²⁶, R²⁶ is a member selected from—CH₃, —CH₂CH₃, —(CH₂)₂CH₃, —CH(CH₃)₂, —CH₂CF₃, —CH₂CHF₂, —CH₂CH₂(OH),—CH₂CH₂(OCH₃), —CH₂CH₂(OC(CH₃)₂), —C(O)CH₃, —CH₂CH₂OC(O)CH₃,—CH₂C(O)OCH₂CH₃, —CH₂C(O)OC(CH₃)₃, —(CH₂)₃C(O)CH₃, —CH₂C(O)OC(CH₃)₃,cyclopentyl, cyclohexyl,

In another embodiment when R^(b) is NR²⁷R²⁸, R²⁷ and R²⁸ are membersindependently selected from H, substituted or unsubstituted alkyl,substituted or unsubstituted heteroalkyl, substituted or unsubstitutedcycloalkyl, substituted or unsubstituted heterocycloalkyl, substitutedor unsubstituted aryl, and substituted or unsubstituted heteroaryl. Inanother embodiment when R^(b) is NR²⁷R²⁸, R²⁷ is H or unsubstitutedalkyl; and R²⁸ is unsubstituted alkyl or alkyl substituted with a memberselected from hydroxyl, phenyl, unsubstituted alkoxy and alkoxysubstituted with a phenyl. In a further embodiment when R^(b) isNR²⁷R²⁸, R²⁷ is H or CH₃.

In another embodiment when R^(b) is NR²⁷R²⁸, R²⁷ and R²⁸ areindependently selected from substituted or unsubstituted alkyl. Inanother embodiment when R^(b) is NR²⁷R²⁸, R²⁷ is unsubstituted alkyl;and R²⁸ is substituted or unsubstituted alkyl. In another embodimentwhen R^(b) is NR²⁷R²⁸, R²⁷ is unsubstituted alkyl; and R²⁸ is alkyl,substituted with a member selected from substituted or unsubstitutedalkoxy and hydroxyl. In another embodiment when R^(b) is NR²⁷R²⁸, R²⁷ isunsubstituted alkyl; and R²⁸ is alkyl, substituted with unsubstitutedalkoxy. In another embodiment when R^(b) is NR²⁷R²⁸, R²⁷ isunsubstituted alkyl; and R²⁸ is alkyl, substituted with alkoxy,substituted with phenyl. In another embodiment when R^(b) is NR²⁷R²⁸,R²⁷ is unsubstituted alkyl; and R²⁸ is alkyl, substituted withunsubstituted alkoxy. In another embodiment when R^(b) is NR²⁷R²⁸, R²⁷and R²⁸ together with the nitrogen to which they are attached, arecombined to form a 4- to 8-membered substituted or unsubstitutedheterocycloalkyl ring. In another embodiment when R^(b) is NR²⁷R²⁸, R²⁷and R²⁸ together with the nitrogen to which they are attached, arecombined to form a 5- or 6-membered substituted or unsubstitutedheterocycloalkyl ring.

In another embodiment, R^(b) is selected from N(CH₃)₂,N(CH₃)(CH₂CH₂(OCH₃)), N(CH₃)(CH₂CH₂OH), NH₂, NHCH₃, NH(CH₂CH₂(OCH₃)),NH(CH₂CH₂(OCH₂Ph), NH(CH₂Ph), NH(C(CH₃)₃) and NH(CH₂CH₂OH). In anotherembodiment, Rb is selected from

Additional antimicrobial compounds are also disclosed previously in U.S.Pat. No. 8,039,450, and patent application publication US 2009/0291917,the contents of which are hereby incorporated by reference in theirentireties.

The practice of the present invention involves the use of one or morecyclopropene compound. As used herein, a cyclopropene compound is anycompound with the formula

where each R¹, R², R³ and R⁴ is independently selected from the groupconsisting of H and a chemical group of the formula:-(L)_(n)-Zwhere n is an integer from 0 to 12. Each L is a bivalent radical.Suitable L groups include, for example, radicals containing one or moreatoms selected from H, B, C, N, O, P, S, Si, or mixtures thereof. Theatoms within an L group may be connected to each other by single bonds,double bonds, triple bonds, or mixtures thereof. Each L group may belinear, branched, cyclic, or a combination thereof. In any one R group(i.e., any one of R¹, R², R³ and R⁴) the total number of heteroatoms(i.e., atoms that are neither H nor C) is from 0 to 6. Independently, inany one R group the total number of non-hydrogen atoms is 50 or less.Each Z is a monovalent radical. Each Z is independently selected fromthe group consisting of hydrogen, halo, cyano, nitro, nitroso, azido,chlorate, bromate, iodate, isocyanato, isocyanido, isothiocyanato,pentafluorothio, and a chemical group G, wherein G is a 3- to14-membered ring system.

The R¹, R², R³, and R⁴ groups are independently selected from thesuitable groups. Among the groups that are suitable for use as one ormore of R¹, R², R³, and R⁴ are, for example, aliphatic groups,aliphatic-oxy groups, alkylphosphonato groups, cycloaliphatic groups,cycloalkylsulfonyl groups, cycloalkylamino groups, heterocyclic groups,aryl groups, heteroaryl groups, halogens, silyl groups, other groups,and mixtures and combinations thereof. Groups that are suitable for useas one or more of R¹, R², R³, and R⁴ may be substituted orunsubstituted.

Among the suitable R¹, R², R³, and R⁴ groups are, for example, aliphaticgroups. Some suitable aliphatic groups include, for example, alkyl,alkenyl, and alkynyl groups. Suitable aliphatic groups may be linear,branched, cyclic, or a combination thereof. Independently, suitablealiphatic groups may be substituted or unsubstituted.

As used herein, a chemical group of interest is said to be “substituted”if one or more hydrogen atoms of the chemical group of interest isreplaced by a substituent.

Also among the suitable R¹, R², R³, and R⁴ groups are, for example,substituted and unsubstituted heterocyclyl groups that are connected tothe cyclopropene compound through an intervening oxy group, amino group,carbonyl group, or sulfonyl group; examples of such R¹, R², R³, and R⁴groups are heterocyclyloxy, heterocyclylcarbonyl, diheterocyclylamino,and diheterocyclylaminosulfonyl.

Also among the suitable R¹, R², R³, and R⁴ groups are, for example,substituted and unsubstituted heterocyclic groups that are connected tothe cyclopropene compound through an intervening oxy group, amino group,carbonyl group, sulfonyl group, thioalkyl group, or aminosulfonyl group;examples of such R¹, R², R³, and R⁴ groups are diheteroarylamino,heteroarylthioalkyl, and diheteroarylaminosulfonyl.

Also among the suitable R¹, R², R³, and R⁴ groups are, for example,hydrogen, fluoro, chloro, bromo, iodo, cyano, nitro, nitroso, azido,chlorato, bromato, iodato, isocyanato, isocyanido, isothiocyanato,pentafluorothio, acetoxy, carboethoxy, cyanato, nitrato, nitrito,perchlorato, allenyl, butylmercapto, diethylphosphonato,dimethylphenylsilyl, isoquinolyl, mercapto, naphthyl, phenoxy, phenyl,piperidino, pyridyl, quinolyl, triethylsilyl, trimethylsilyl, andsubstituted analogs thereof.

As used herein, the chemical group G is a 3- to 14-membered ring system.Ring systems suitable as chemical group G may be substituted orunsubstituted; they may be aromatic (including, for example, phenyl andnapthyl) or aliphatic (including unsaturated aliphatic, partiallysaturated aliphatic, or saturated aliphatic); and they may becarbocyclic or heterocyclic. Among heterocyclic G groups, some suitableheteroatoms are, for example, nitrogen, sulfur, oxygen, and combinationsthereof. Ring systems suitable as chemical group G may be monocyclic,bicyclic, tricyclic, polycyclic, spiro, or fused; among suitablechemical group G ring systems that are bicyclic, tricyclic, or fused,the various rings in a single chemical group G may be all the same typeor may be of two or more types (for example, an aromatic ring may befused with an aliphatic ring).

In one embodiment, one or more of R¹, R², R³, and R⁴ is hydrogen or(C₁-C₁₀) alkyl. In another embodiment, each of R¹, R², R³, and R⁴ ishydrogen or (C₁-C₈) alkyl. In another embodiment, each of R¹, R², R³,and R⁴ is hydrogen or (C₁-C₄) alkyl. In another embodiment, each of R¹,R², R³, and R⁴ is hydrogen or methyl. In another embodiment, R¹ is(C₁-C₄) alkyl and each of R², R³, and R⁴ is hydrogen. In anotherembodiment, R¹ is methyl and each of R², R³, and R⁴ is hydrogen, and thecyclopropene compound is known herein as 1-methylcyclopropene or“1-MCP.”

In another embodiment, the cyclopropene is of the formula:

wherein R is a substituted or unsubstituted alkyl, alkenyl, alkynyl,cycloalkyl, cycloalkylalkyl, phenyl, or naphthyl group; wherein thesubstituents are independently halogen, alkoxy, or substituted orunsubstituted phenoxy. In one embodiment, R is C₁-C₈ alkyl. In anotherembodiment, R is methyl.

In another embodiment, the cyclopropene is of the formula:

wherein R¹ is a substituted or unsubstituted C₁-C₄ alkyl, C₁-C₄ alkenyl,C₁-C₄ alkynyl, C₁-C₄ cycloalkyl, cycloalkylalkyl, phenyl, or napthylgroup; and R², R³, and R⁴ are hydrogen. In another embodiment, thecyclopropene comprises 1-methylcyclopropene (1-MCP).

As used herein, the phrase “transgene vector” refers to a vector thatcontains an inserted segment of deoxyribonucleic acid (DNA), the“transgene” that is transcribed into messenger ribonucleic acid (mRNA)or replicated as ribonucleic acid (RNA) within a host cell. The phrase“transgene” refers not only to that portion of inserted DNA that isconverted into RNA, but also those portions of the vector that arenecessary for the transcription or replication of the RNA. A transgenetypically comprises a gene-of-interest but needs not necessarilycomprise a polynucleotide sequence that contains an open reading framecapable of producing a protein.

Meats, plants, or plant parts may be treated in the practice of thepresent invention. One example is treatment of whole plants; anotherexample is treatment of whole plants while they are planted in soil,prior to the harvesting of useful plant parts.

Any plants that provide useful plant parts may be treated in thepractice of the present invention. Examples include plants that providefruits, vegetables, and grains.

As used herein, the phrase “plant” includes dicotyledonous plants andmonocotyledonous plants. Examples of dicotyledonous plants includetobacco, Arabidopsis, soybean, tomato, papaya, canola, sunflower,cotton, alfalfa, potato, grapevine, pigeon pea, pea, Brassica, chickpea,sugar beet, rapeseed, watermelon, melon, pepper, peanut, pumpkin,radish, spinach, squash, broccoli, cabbage, carrot, cauliflower, celery,Chinese cabbage, cucumber, eggplant, and lettuce. Examples ofmonocotyledonous plants include corn, rice, wheat, sugarcane, barley,rye, sorghum, orchids, bamboo, banana, cattails, lilies, oat, onion,millet, and triticale. Examples of fruit include banana, pineapple,oranges, grapes, grapefruit, watermelon, melon, apples, peaches, pears,kiwifruit, mango, nectarines, guava, persimmon, avocado, lemon, fig, andberries.

Those skilled in the art would understand certain variation can existbased on the disclosure provided. Thus, the following examples are givenfor the purpose of illustrating the invention and shall not be construedas being a limitation on the scope of the invention or claims.

EXAMPLES Example 1

12-Well (7 milliliter (mL volume per well) microtiter plates are usedfor the in vitro inhibition assay for volatile antimicrobial compounds.A 3-mL volume of full-strength Potato Dextrose Agar (PDA) is added toeach well. After cooling, 1 microliter (μL) of 1×10⁶ per mL Botrytiscinerea spore suspension is spot pipetted to the center of the agar. Forthe first experiment, inoculated plates are allowed to germinate for 5days at 4° C. For the second experiment, plates are inoculatedimmediately prior to volatile fungicide treatment. Small Whatman #1filter disks (Cat. No. 1001-0155) are placed, in duplicate, on theunderside of a polyethylene PCR plate sealing film.

TABLE 1 Results of in vitro assay for volatile fungicide Rate ofCompound A (mg per disk) Botrytis inhibition % (in vitro) 1.25 100% 0.63100% 0.31 100% 0.16 100% 0.08 100% 0.04 100% 0.023 100% 0.01 100% 0.005100% 0.0024 85% 0.001 69% 0.0006 46% Control 0%

For determination of the minimum inhibitory concentration (MIC),Compound A (benzoxaborole; FIG. 1) is diluted in acetone, and theappropriate amount of compound is added to disks in a dose dependentmanner (1.25 to 0.0006 milligrams per disk (mg/disk)). The acetone ispermitted to evaporate for 5 minutes. The headspace around the Botrytiscinerea inoculum is then sealed inside the well by the film with theadhering disk containing the fungicide. Plates are inverted, placed overthe treated disks and sealed to prevent any of the chemical from flakingfrom the disk and falling onto the inoculated agar. After 14 days ofstorage at 4° C., cultures are evaluated for percent growth relative tocontrol. Regardless of whether the spores had germinated for 5 days, orif the treatment commenced soon after inoculation of the plates (˜15minutes); there is 100% control of the fungal pathogen down to 0.005 mg.Experimental results are summarized in Table 1. The results suggest thatCompound A is able to kill Botrytis cinerea spores and inhibit mycelialgrowth at the same concentration. Thus, Compound A (FIG. 1) shows 100%efficacy in the in vitro inhibition of fungal growth at a rate of 0.005mg/disk.

Example 2

A total of 14 antimicrobial compounds are tested using the in vitroinhibition assay described in Example 1. All 14 compounds are applied tothe Whatman disks, in duplicate, in a dose dependent manner (0.31 to0.0006 mg/disk). The results show that Compound A provides the bestcontrol of Botrytis cinerea, with 100% control down to 0.005 mg/disk.Other compounds, such as Compound C, Compound D, and Compound Econferred 100% control down to 0.023, 0.04, and 0.08 mg/disk,respectively. The tested compounds are shown in FIG. 3. Results of ninecompounds are summarized in Table 2, where the other five compounds showno detected activity in the ranges tested.

TABLE 2 Results of in vitro assay for volatile fungicide in % Botrytisinhibition Rate Comp. Comp. Comp. Comp. Comp. Comp. Comp. Comp. Comp.(mg/disk) A C D E F G H J K 0.31 100% 100% 100% 100% 70% 100% 85% 50%48% 0.16 100% 100% 100% 100% 53%  78% 80% 13% 29% 0.08 100% 100% 100%100% 40%  43% 55%  8%  5% 0.04 100% 100% 100%  79% 18%  13% 38%  5%  0%0.023 100% 100%  80%  79% 10%  3% 18%  0%  0% 0.01 100%  83%  70%  69% 8%  0%  3%  0%  0% 0.005 100%  63%  38%  38%  8%  0%  0%  0%  0% 0.0024 85%  43%  15%  28%  0%  0%  0%  0%  0% 0.001  69%  15%  0%  13%  0%  0% 0%  0%  0% 0.0006  46%  0%  0%  13%  0%  0%  0%  0%  0% Control  0%  0% 0%  0%  0%  0%  0%  0%  0%

Example 3

Compound B (FIG. 2; 2-(hydroxymethyl)phenylboronic acid cyclicmonoester, a des-fluoro analogue of Compound A), is evaluated in asimilar manner as described in Examples 1 and 2 above. The compound isapplied to the Whatman filter paper at rates from 0.5 mg to 0.0039mg/disk. Results show that Compound B inhibits 100% Botrytis cinerea ata rate of 0.0078 mg/disk.

Example 4

In order to assess the in vivo activity of volatile antimicrobialcompounds, a volatile bioassay is developed using green table grape.Fruit are placed individually inside a 20 mL scintillation vial, withthe stem wound facing upwards. The fresh stem wound is inoculated with10 μL of 1×10⁶ per mL Botrytis cinerea spore suspension. Whatman filterpaper (Cat. No. 1822-024) is placed inside duplicate vial caps. Fordetermination of the MIC, Compound A (FIG. 1) is diluted in acetone, andthe appropriate amount of compound is added to the disks in a dosedependent manner (2.5 to 0.0024 mg/disk). The acetone is permitted toevaporate for 5 minutes. The vials are then capped with the lidscontaining the fungicide, and placed for 14 days at 4° C. After storage,fruit are evaluated for incidence of disease and appearance ofphytotoxicity. Results are summarized in Table 3 and there is 100%control of Botrytis cinerea down to 0.04 mg/disk and no evidence ofphytotoxicity at any of the rates evaluated. Representative photos ofexemplary in vivo inhibition results using Compound A are shown in FIGS.4A-4D, where 0.04 mg of Compound A shows 100% inhibition and 0.0024 mgof Compound A shows no inhibition.

TABLE 3 Results of in vivo assay for volatile fungicide Rate of CompoundA (mg per disk) Botrytis inhibition % (in vivo) 1.25 100% 0.63 100% 0.31100% 0.16 100% 0.08 100% 0.04 100% 0.023 0% 0.01 0% 0.005 0% 0.0024 0%Control 0%

Example 5

In order to assess the in vivo activity of volatile antimicrobialcompounds, a volatile bioassay is developed using strawberry. Two fruitare placed inside a 240 mL jar, with the calyx facing downwards. A freshwound is inoculated with 20 μL of 1×10⁶ per mL Botrytis cinerea sporesuspension. Whatman filter paper (Cat. No. 1822-024) is placed insideduplicate jar lids. For determination of the MIC, Compound A(benzoxaborole; FIG. 1) is diluted in acetone, and the appropriateamount of compound is added to the disks in a dose dependent manner (2.5to 0.005 mg/disk). For determination of the MIC, Compound B(benzoxaborole; FIG. 2) is diluted in acetone, and the appropriateamount of compound is added to the disks in a dose dependent manner (2.5to 0.005 mg/disk). The acetone is permitted to evaporate for 5 minutes.The jars are then capped with the lids containing the fungicide, andplaced for 5 days at 21° C. After storage, fruit are evaluated forincidence and severity of disease and appearance of phytotoxicity.Results are summarized in Table 4. There is 100% control of Botrytiscinerea down to 0.16 mg/disk for Compound A and 100% control of Botrytiscinerea down to 0.32 mg/disk for Compound B, and no evidence ofphytotoxicity at any of the rates evaluated.

TABLE 4 Percent (%) Incidence of Botrytis cinerea on Strawberry (invivo) Rates (mg/disk) Compound A Compound B 0.005 75% 100% 0.01 100%100% 0.02 50% 100% 0.04 75% 75% 0.08 0% 50% 0.16 0% 25% 0.32 0% 0% 0.640% 0% 1.25 0% 0% 2.5 0% 0%

Example 6

In order to assess the in vivo dose by time activity of volatileantimicrobial compounds, a volatile bioassay is developed usingstrawberry. Two fruit are placed inside a 240 mL jar, with the calyxfacing downwards. A fresh wound is inoculated with 20 μL of 1×10⁶ per mlBotrytis cinerea spore suspension. Whatman filter paper (Cat. No.1822-024) is placed inside duplicate jar lids. Compound A(benzoxaborole; FIG. 1) is diluted in acetone, and the appropriateamount of compound is added to the disks at two rates 0.008 or 0.125 mg.The acetone is permitted to evaporate for 5 minutes. The jars are cappedwith the lids containing the fungicide, and incubated with the volatilefungicide for 1, 3, 6, 24 or 72 hours. After incubation, lids containingthe disk with Compound A are replaced with new lids without Compound A.All samples are maintained at 21° C. for 3 days, and then the lids areremoved and maintained for an additional 48 hours, all at 90% relativehumidity (R.H.). The fruit are evaluated for incidence and severity ofdisease and appearance of phytotoxicity. Results are summarized in Table5. There is 100% control of Botrytis cinerea at 0.125 mg/disk forCompound A after 6 hour exposure, and no evidence of phytotoxicity.0.125 mg of Compound A shows 100% in vivo inhibition in comparison tothe acetone only control. A representative result is also shown in FIGS.5A and 5B.

TABLE 5 Incidence (%) and Severity of Botrytis cinerea on strawberriesover time Compound A Incidence (%) Severity (0 to 3) Rates (mg/disk)Time (h) 0.008 0.125 0.008 0.125 1 100% 67% 4.0 2.3 3 0% 33% 0.0 1.3 633% 0% 1.0 0.0 24 67% 0% 2.3 0.0 72 33% 0% 1.0 0.0 Severity: 0 = nofungal growth 1 = slight infection (<5 millimeter (mm) diameter) 2 =moderate infection (<1 centimeter (cm) diameter) 3 = high infection (>1cm diameter) 4 = extreme infection (> half-length of fruit)

Example 7

12-Well (7 mL volume per well) microtiter plates are used for the invitro inhibition assay for volatile antimicrobial compounds. A 3-mLvolume of full-strength LB Agar is added to each well. After cooling, 15μL of Escherichia coli, adjusted to an optical density of 0.02 to 0.035,and further diluted 1/10 is pipetted to the center of the agar andtilted to distribute uniformly. Small Whatman #1 filter disks (Cat. No.1001-0155) are placed, in duplicate, on the underside of a polyethylenepolymerase chain reaction (PCR) plate sealing film. For determination ofthe minimum inhibitory concentration (MIC), Compound A (benzoxaborole;FIG. 1) is diluted in acetone, and 5 mg of compound is added to thedisks. The acetone is permitted to evaporate for 5 minutes. Theheadspace around the Escherichia coli inoculum is then sealed inside thewell by the film with the adhering disk containing the fungicide. Platesare inverted, placed over the treated disks and sealed to prevent any ofthe chemical from flaking from the disk and falling onto the inoculatedagar. After 3 days of storage at 4° C., cultures were transferred to 23°C. for an additional 2 days, and then evaluated for colony growthrelative to control. Experimental results are summarized in

TABLE 6 The results suggest that Compound A is able to inhibitEscherichia coli. Table 6. Results of in vitro assay for volatilefungicide Rate of Compound A (mg per disk) Colony Rating 5.00 1Untreated 3 Not Inoculated 0 Colony Rating: 0 = No colonies 1 = Microcolonies not connected 2 = Small colonies with some merging 3 = Largecolonies merging together

Example 8

12-Well (6.5 mL volume per well) microtiter plates are used for the invitro inhibition assay for volatile antimicrobial compounds. A 3-mLvolume of full-strength Potato Dextrose Agar (PDA) is added to eachwell. After cooling, 1 μL of 1×10⁵ per mL Botrytis cinerea, Penicilliumexpansum, Alternaria alternata, Monilinia fructicola or Glomerellacingulata spore suspension is spot-pipetted to the center of the agar.Plates are inoculated immediately prior to volatile fungicide treatment.A Whatman #1 filter disk (Cat. No. 1001-0155) is placed, in duplicate,on the underside of a polyethylene PCR plate sealing film. Fordetermination of the minimum inhibitory concentration (MIC), compoundsare diluted in acetone, and the appropriate amount of compound is addedto the disks in a dose dependent manner to achieve a final headspaceconcentration of 1142.9 to 0.6 mg/L. The acetone is permitted toevaporate for 5 minutes. The headspace around the inoculum is thensealed inside the well by the film with the adhering disk containing thefungicide by inverting the plates over the treated disks and sealing toprevent any of the chemical from flaking from the disk and falling ontothe inoculated agar. After 3 days of storage at 23° C., the cultures areevaluated for percent growth relative to control based on measurement offungal colony diameter. Experimental results are summarized in Table 7.The results indicate that benzoxaborole compounds have excellent invitro activity against five selected plant fungal pathogens.

TABLE 7 MIC (mg/L, headspace concentration) of numerous benzoxaborolecompounds applied as a volatile treatment against numerous plant fungalpathogens (Compound 10 is the same as Compound A, and Compound 11 is thesame as Compound B). Cmpd MIC (mg/L) Structure # BOTRCI PENIEX ALTEALMONIFC GLOMCI

6 2.2 17.9 4.5 8.9 17.9

7 2.2 17.9 8.9 8.9 71.4

8 2.2 35.7 8.9 4.5 71.4

9 2.2 8.9 8.9 8.9 35.7

10 2.2 2.2 <0.6 <0.6 <0.6

11 4.5 17.9 4.5 2.2 35.7

30 2.2 8.9 2.2 2.2 n/a

34 <0.6 2.2 2.2 n/a n/a

200 10.6 68.3 7.3 6.3 n/a

201 3.8 29.5 16.1 8.5 9.3 BOTRCI = Botrytis cinerea (gray mold) PENIEX =Penicillium expansum (blue mold of apple) ALTEAL = Alternaria alternata(brown spot of tobacco) MONIFC = Monilinia fructicola (brown rot ofapple) GLOMCI = Glomerella cingulata (anthracnose of pepper)

Example 9

12-Well (6.5 mL volume per well) microtiter plates are used for the invitro inhibition assay for volatile antimicrobial compounds. A 3-mLvolume of full-strength Potato Dextrose Agar (PDA) is added to eachwell. After cooling, 1 μL of 1×10⁵ per mL Botrytis cinerea andPenicillium expansum spore suspension is spot-pipetted to the center ofthe agar. Plates are inoculated immediately prior to volatile fungicidetreatment. A Whatman #1 filter disk (Cat. No. 1001-0155) is placed, induplicate, on the underside of a polyethylene PCR plate sealing film.For determination of the minimum inhibitory concentration (MIC),compounds are diluted in acetone, and the appropriate amount of compoundis added to the disks in a dose dependent manner to achieve a finalheadspace concentration of 35.7 to 0.03 mg/L. The acetone is permittedto evaporate for 5 minutes. The headspace around the inoculum is thensealed inside the well by the film with the adhering disk containing thefungicide by inverting the plates over the treated disks and sealing toprevent any of the chemical from flaking from the disk and falling ontothe inoculated agar. After 3 days of storage at 23° C., the cultures areevaluated for percent growth relative to control based on measurement offungal colony diameter. Experimental results are summarized in Table 8.The results indicate that numerous benzoxaborole compounds haveexcellent in vitro activity against two selected plant fungal pathogens.

TABLE 8 MIC (mg/L) of numerous benzoxaborole compounds applied as avolatile treatment against Botrytis cinerea and Penicillium expansumplant fungal pathogens. Cmpd MIC (mg/L) Structure # BOTRCI PENIEX

21 1.1 35.7

22 4.5 35.7

38 0.6 8.9

39 0.6 8.9

54 0.6 4.5

55 4.5 >35.7

62 2.2 8.9

63 1.1 17.9

64 1.1 8.9

72 35.7 >35.7

73 35.7 >35.7

74 2.2 35.7

86 0.6 8.9

87 0.6 8.9

105 0.6 4.5

114 17.9 >35.7

115 0.6 8.9

116 1.1 8.9

121 4.5 17.9

122 2.2 17.9

124 4.5 8.9

127 2.2 4.5

129 4.5 8.9

130 1.1 4.5

132 1.1 4.5

133 8.9 35.7

134 17.9 >35.7

135 17.9 >35.7

136 8.9 >35.7

137 0.3 1.1

202 35.7 142.9

203 8.9 142.9

204 8.9 >35.7 BOTRCI = Botrytis cinerea (gray mold) PENIEX = Penicilliumexpansum (blue mold of apple)

Example 10

12-Well (6.5 mL volume per well) microtiter plates are used for the invitro inhibition assay for volatile antimicrobial compounds A and B(FIG. 1) against numerous plant fungal pathogens. A 3-mL volume offull-strength Potato Dextrose Agar (PDA) is added to each well. Aftercooling, 1 μL of 1×10⁵ spores per mL of Botrytis cinerea, Penicilliumexpansum, Alternaria alternata, Glomerella cingulata, Penicilliumdigitatum, Monilinia fruticola, Aspergillus brasiliensis, Colletotrichumacutatum, Fusarium sambucinum, Phytophthora capsici, Geotrichumcandidum, Aspergillus niger, Diplodia gossypina or Diaporthe citriisuspension is spotted onto the center of the agar. A Whatman #1 filterdisk (Cat. No. 1001-0155) is placed, in duplicate, on the underside of apolyethylene PCR plate sealing film. For determination of the minimuminhibitory concentration (MIC), test compounds are diluted in acetone,and the appropriate amount of compound is added to the disks in a dosedependent manner to achieve a final headspace concentration of 35.7 to0.03 mg/L. The acetone is permitted to evaporate for five minutes. Theheadspace around the inoculum is then sealed inside the well by the filmwith the adhering disk containing the fungicide by inverting the platesover the treated disks and sealing to prevent any of the chemical fromflaking from the disk and falling onto the inoculated agar. After 3 daysof storage at 23° C., cultures are evaluated for percent growth relativeto control. Results shown in Table 9 demonstrate the ability ofbenzoxaborole compounds A and B to control the growth of numerous fungalplant pathogens through volatile activity.

TABLE 9 MIC (mg/L) of Compounds A and B applied as a volatile againstnumerous fungal plant pathogens Compound A Compound B Pathogens MIC MICB. cinerea 2.2 4.5 P. expansum 1.1 8.9 M. fruticola 2.2 1.1 A. alternata2.2 2.2 G. cingulata 17.9 35.7 P. digitatum 2.2 4.5 A. brasiliensis 2.20.6 C. acutatum 4.4 8.9 F. sambucinum 1.1 4.5 P. capsici 1.1 n/a G.candidum 8.9 8.9 A. niger 2.2 1.1 M. piriformis 1.1 2.2 D. gossypina 1.14.5 D. citrii 2.2 17.9

Example 11

12-Well (6.5 mL volume per well) microtiter plates are used for the invitro inhibition assay for volatile antimicrobial Compound A (FIG. 1)against numerous bacterial pathogens. A 3-mL volume of Nutrient agar isadded to each well and allowed to dry before introducing the pathogen.Escherichia coli, Pectobacterium carotovorum, Xanthomonas axonopodis andSalmonella enterica cell suspensions are adjusted to an optical densityof 0.2 to 0.35, and further diluted 1/10, and 15 μL is pipetted to thecenter of each well and tilted to distribute uniformly. A Whatman #1filter paper (CAT 1001-0155) is placed on the underside of apolyethylene PCR plate sealing film. For determination of minimumbactericidal concentration (MBC), Compound A is diluted in acetone, and50 μL are applied to the disks, in duplicate, in a dose dependent mannerin order to achieve a final headspace concentration of 71.4 to 0.03mg/L. The acetone is permitted to evaporate for 5 minutes. The filmswith the treated disks are then applied over the inoculated plates andsealed. Plates are inverted, and incubated at 23° C. for 48 hours. Afterthe incubation period, the bacteria colonies are dislodged in sterilewater containing tween 80 (0.001%) and the optical density (OD; 600 nm)is determined Results are summarized in Table 10, where the headspaceconcentration required to control at least 80% of bacterial growth isreported. Compound A shows good antimicrobial activity against numerousbacteria in this in vitro assay.

TABLE 10 Rate (mg/L) of Compound A offering at least 80% control againstbacterial pathogens E. coli P. carotovorum X. axonopodis S. enterica35.7 2.2 4.5 17.9

Example 12

In order to assess the in vivo activity of volatile antimicrobialCompound A, a volatile bioassay is developed to evaluate the control ofEscherichia coli and Salmonella enterica on fresh beef. The beef iswashed to remove any natural inoculum by rinsing in warm water for 2minutes. Two strips, single layer, are placed in a sterile 10.8-cupSnapWare airtight container (Model #109842).

TABLE 11 Colony forming unit (CFU/mL) and log reductions of E. coli andS. enterica from beef after a volatile treatment with Compound A. LogPathogens Treatments Log CFU/mL reduction E. coli Control 8.27 3.17Compound A 5.09 S. enterica Control 7.38 2.43 Compound A 4.95

Each strip is inoculated on the surface by placing 20 μL of either E.coli or S. enterica cell suspensions that are adjusted to an opticaldensity of 0.35 (600 nm), and further diluted 1/10. For determination ofefficacy, Compound A powder is introduced into the container with asublimation device (copper tube heated to 200° C. with fan flow at 0.5liters per minute (L/min)) at a rate required to achieve a finalheadspace concentration of 100 mg/L. The containers and their contentsare then incubated for 2 days at 21° C. After treatment, the beef iswashed, and the wash is collected, serially diluted, plated on nutrientagar, and then incubated for an additional 24 hours at 37° C. Bacterialcolonies are counted and expressed as colony forming units (CFU/mL),with the log reduction calculated relative to the control. Resultslisted in Table 11 show good antimicrobial activity of Compound Aagainst E. coli and S. enterica in this in vivo assay using beef.Compound A demonstrates a 3.17 log reduction (>99.9%) of E. coli and a2-log reduction for S. enterica.

Example 13

In order to assess the in vivo activity of volatile antimicrobialCompound A on controlling Botrytis cinerea on ornamental flowers, avolatile bioassay is developed using white carnations.

TABLE 12 Botrytis cinerea incidence on carnations treated with CompoundA Compound A Disease incidence (%) on Petals Rates (mg/L) Day 0 Day 1Day 2 Day 3 Day 8 1 0 0 0 4 16 0.2 0 8 20 16 36 0.04 0 0 16 40 92Control 0 68 92 96 100

Five carnations are placed in an 800 mL jar containing 200 mL of acommon commercial flower preservative. Five jars are then placed in a117 L Rubbermaid storage box (Cat #2244). The petals are uniformlyspray-inoculated with 5 mL of 1×10⁵ spores/mL of Botrytis cinereasuspension. The tub is closed tightly. For treatment application,Compound A is dissolved in an aqueous 1,2-propylene glycol solution(3:1) and 5 mL of the solution is volatilized into the container usingan ES-100-H SmartFog system (Reno, Nev.) through a ½″ side port that issealed immediately after the application. The flowers are incubated for3 days at 21° C. After storage, the flowers are evaluated for incidencebased on presence of disease on flower petals relative to untreatedcontrol flowers for up to 8 days at 21° C., with results summarized inTable 12. Compound A at 1 mg/L shows 0% incidence 2 days after treatmentremoval and only 16% incidence after 8 days, and generally demonstratesgood volatile antimicrobial activity against Botrytis cinerea in this invivo analysis of infection in an ornamental flower.

Example 14

A similar test like the one described above is also performed on whitecarnations (treated with or without the commercial anti-ethylenecompound silver thiosulfate; STS) with natural inoculum. Compound A iseither dissolved in an aqueous 1,2-propylene glycol solution (3:1) and 5mL of the solution volatilized using an ES-100-H SmartFog system (Reno,Nev.) through a ½″ side port that is sealed immediately after theapplication, or dissolved in acetone and applied to a 42.5 millimeter(mm) Whatman #1 filter disk (Cat. No. 1001-042), and placed on a watchglass after allowing the acetone to evaporate for 5 minutes. The flowersare incubated for 3 days at 21° C. After storage, the flowers areevaluated for an additional 8 days for disease severity based on thenumber of lesions present on flower petals and sepals. Results listed inTable 13 show good antimicrobial activity against Botrytis cinerea inthis in vivo analysis.

TABLE 13 Botrytis cinerea severity after 8 days of shelf-life based onnumber of lesions on petals and sepals after an active fog or passivevolatile treatment with Compound A. Severity (Average Number of Lesions)Compound A Non-STS STS Plant Part Rate (mg/L) Fog Volatile Fog VolatilePetals 1 0 1.5 0 0.1 0.2 0 1.8 0 0.1 0.04 0.4 3.1 0.3 0.5 0 2.1 18.8 7.743.2 Sepals 1 0.04 1 0.04 0.2 0.2 0.04 1.6 0.1 0.3 0.04 0.3 2.1 0.6 1.10 4.5 4.8 6.8 3.6

Example 15

A similar test like the one described above is also performed on whiteroses with natural inoculum. Five white roses are placed in an 800 mLjar containing 200 mL of a common commercial flower preservative.

TABLE 14 Botrytis cinerea incidence and severity based on infection onpetals and sepals of white roses after a three day volatile treatment ofCompound A at 21° C., and an additional two days at 21° C. Appliedthrough Volatilized from sublimation Whatman Filter Compound A IncidenceIncidence Severity Rate (mg/L) (%) Severity (0-4)* (%) (0-4)* 1 0 0 53.31.6 0.2 13.3 0.5 66.7 1.8 0.04 46.7 1.7 46.7 1.1 Control- Acetone 80 2.986.7 2.4 Control 100 3.1 100 3.1 *Severity Rating 0 = No disease 1 =Browning and small lesions on the sepals or petals 2 = Browning, petalscovered with fungal spores 3 = Browning, petals covered with fungalspores, some petal drop 4 = Browning, petals covered with fungal spores,some flowers aborted

Three jars are then placed in a 117 L Rubbermaid storage box (Cat#2244). Two small fans are place in opposite ends of the container toassist with the volatile distribution of compound A. The tub is closedtightly, and then Compound A is diluted in acetone, and then pipettedonto a 1.5 inch×1 inch cotton strip. The acetone is allowed to evaporatefor five minutes. Compound A is then introduced to the containers by asublimation device (copper tube heated to 200° C. with fan flow at 0.5L/min) to achieve a final headspace concentration of 0.04, 0.2, 1 mg/L,through a ½″ side port that is sealed immediately after the application.Alternatively, Compound A is pipetted onto a 42.5 mm Whatman #1 filterdisk (Cat. No. 1001-042), supported by a watch glass, where the acetoneis allowed to evaporate for five minutes prior to sealing the container.The flowers are incubated for three days at 21° C. After treatment,flowers are evaluated for an additional two days for disease incidenceand severity of the flower petals. Applying a treatment at 1 mg/Lthrough sublimation results in 0% incidence. Rose petals after treatmentwith Compound A have no disease incidence, retained white color, and theroses had no petal drop. Results listed in Table 14 show goodantimicrobial activity against Botrytis cinerea infection of whiteroses, and that enhancing the rate of volatilization through sublimationresulted in greater disease control.

Example 16

To test the effect of Compound A (FIG. 1) on vegetables, potato, onionand squash were obtained from a local store, and the surface sterilizedwith 0.825% sodium hypochlorite (NaOCl). A slice of potato or two leavesof onion were placed in a sterile Petri plate, while whole squash wereplaced in a sterile 10.8-cup SnapWare airtight container (Model#109842). Each slice of potato was inoculated with 20 μL of a 1×10⁵spores/mL Fusarium sambucinum suspension, while onions were inoculatedwith 20 μL of a 1×10⁶ spores/mL Botrytis cinerea suspension. Forinoculation of squash, a small core was removed, and a mycelial plug ofPhytophthora capsici was inserted and capped with the core. Compound Awas diluted in acetone, and added to a 42.5 mm Whatman #1 filter disk(Cat. No. 1001-042) attached to the inner side of the lid at a rate toachieve a final headspace concentration of 10 mg/L. The acetone waspermitted to evaporate for 5 minutes before parafilm sealing the platesor closing the airtight containers. The vegetables were incubated at 21°C. for 3 days, and evaluated for mycelial growth, dry rot, andwater-soaked appearance (mm diameter) with results summarized in Table13. Compound A demonstrated good fungal control of 3 plant pathogensusing 3 different vegetable crops in this in vivo assay.

TABLE 15 Effect of Compound A at controlling fungal growth on potato,onion and squash. Potato Onion Squash Fusarium Botrytis Phytophthorasambucinum cinerea capsici Mycelial Water Water Mycelial Treatmentsgrowth Dry rot soaked soaked growth 10 mg/L 0 0 0 5.3 1.1Control-acetone 4.3 4.7 7 24 17.4 Control-no acetone 31.6 10.3 8.5 30.924.7

Example 17

To test the effect of Compound A at controlling bacterial pathogens ofvegetables, potato, onion and carrot are chopped into small cubes andsurface sterilized with 0.825% NaOCl and allowed to dry. Four smallcubes (approximately 1 square centimeter (cm²)) of each vegetable areplaced in a sterile Petri plate. Each cube is inoculated with 25 μL ofPectobacterium carotovorum (bacterial concentration of OD 1.0, 600 nm).For determination of efficacy, Compound A is diluted in acetone, and theappropriate volume to achieve a final headspace concentration of 50 mg/Lis added to a 42.5 mm Whatman #1 filter disk (Cat. No. 1001-042)attached to the inner side of the lid. The acetone is permitted toevaporate for five minutes before closing the plate and sealing it withparafilm. The vegetables are incubated at 10 C for four days. Resultslisted in Table 16 demonstrate antimicrobial activity against P.carotovorum on onion (2.14 log reduction), carrot (0.29 log reduction)and potato (0.84 log reduction) in this in vivo analysis.

TABLE 16 Effects of Compound A (50 mg/L) in reducing growth of P.carotovorum on potato, onion and carrot. Crops Treatments Log CFU/mL Logreduction Potato Control 7.47 Compound A 6.63 0.84 Onion Control 8.13Compound A 5.99 2.14 Carrot Control 6.36 Compound A 6.06 0.29

Example 18

In order to assess the in vivo activity of volatile antimicrobialCompound A in fruit, a volatile bioassay is developed using strawberry,grape and blueberry. Eight strawberries, 16 grapes or 30 blueberries(per replicate) are placed in a commercially relevant sized PETclamshell, with the stem end facing up for blueberries and grapes, anddownwards for strawberries. A fresh wound is inoculated with 20 μL(strawberry and grape) or 10 μL (blueberry) of 1×10⁶ per mL Botrytiscinerea spore suspension. The clamshells are placed inside a 10.8-cupSnapWare airtight container (Model #109842). A 42.5 mm Whatman #1 filterdisk (Cat. No. 1001-042) is placed on a watch glass. Compound A isdissolved in acetone and added to the disks in a dose dependent mannerto produce a final headspace concentration of 0.4, 2, or 10 mg/L. Theacetone is permitted to evaporate for five minutes. The containers arethen closed with lids and placed for three days at 21° C. After storage,fruits are evaluated for incidence and severity (0 to 4) of disease foran additional three days at 21° C., with results summarized in Table 17.Results demonstrate good in vivo volatile antimicrobial control ofBotrytis cinerea with approximately 50% lower incidence and dramaticallylower severity for strawberry, grape and blueberry after three days ofshelf-life.

TABLE 17 Effect of a three day volatile treatment of Compound A (0.4, 2or 10 mg/L) in controlling the incidence and severity of B. cinereainfection of strawberry, grape and blueberry during a 3three daypost-treatment evaluation period at 21° C. Strawberry Grape BlueberryCompound A Evaluation Incidence Severity Incidence Severity IncidenceSeverity Rate (mg/L) Days (%) (0-4) (%) (0-4) (%) (0-4) 10 0 7.1 0 0 012.9 0.1 2 0 14.3 0.1 0 0 9.7 0 0.4 0 0 0 3.1 0 21 0.1 Control 0 50 0.4100 2.3 95.2 1.2 10 1 35.7 0.2 0 0 12.9 0.1 2 1 50 0.3 0 0 9.7 0 0.4 121.4 0.1 3.1 0 21 0.2 Control 1 100 1 100 2.5 100 1.7 10 2 42.9 0.5 3.10 12.9 0.2 2 2 50 0.3 0 0 9.7 0.1 0.4 2 21.4 0.1 15.6 0.2 40.3 0.5Control 2 100 2.2 100 2.7 100 1.9 10 3 42.9 0.8 56.3 0.4 41.9 0.6 2 364.3 0.5 56.3 0.3 40.3 0.6 0.4 3 28.6 0.5 62.5 0.5 62.9 1 Control 3 1002.7 100 3.8 100 2.1 *Severity 0 = no fungal growth 1 = slight infection(only visible inside wound with microscope) 2 = moderate infection(visible growth at the point of inoculation) 3 = high infection (>1 cmdiameter cone of Botrytis) 4 = extreme infection (>half-length of fruit)

Example 19

In order to assess the in vivo activity of volatile antimicrobialCompound A in fruit, a volatile bioassay is developed using orangefruit. Two oranges are placed inside a PET clamshell. Three fresh woundsper orange are inoculated with 30 μL of 1×10⁶ per mL Penicilliumdigitatum spore suspension. The clamshells are placed inside a 10.8-cupSnapWare airtight container (Model #109842). A 42.5 mm Whatman #1 filterdisk (Cat. No. 1001-042) is placed on a watch glass. Compound A isdissolved in acetone and added to the disks in a dose dependent mannerto produce a final headspace concentration of 2, 10, or 50 mg/L. Theacetone is permitted to evaporate for five minutes. The containers arethen closed with the lids and placed for three days at 21° C. Afterstorage, fruits are evaluated for disease incidence (mm diameter of therot) and pathogen sporulation (mm diameter) on the surface of the fruitsfor an additional two days at 21° C., with results summarized in Table18. Results demonstrate good in vivo volatile antimicrobial control ofP. digitatum in inoculated orange, especially at rates greater than 10mg/L.

TABLE 18 Incidence and severity of Penicillium digitatum on oranges asdepicted by water soaked lesion and fungal spores on the surface of thefruits Compound A Water soaked Lesions (mm) Sporulation (mm) Rate (mg/L)Day 0 Day 1 Day 2 Day 0 Day 1 Day 2 50 0  0  5  0   0   1.2 10 0  0  9 0.5  0.4  2.5 2 0  0  13.4 0    0.4  2.7 Control 17.8 31.2 52.4 5   15.135.6

Example 20

In order to assess the in vivo activity of volatile antimicrobialCompound A in fruit, a volatile bioassay is developed using apple. Twoapples are placed inside a PET clamshell. Three fresh wounds per appleare inoculated with 30 μL of 1×10⁶ per mL Penicillium expansum sporesuspension. The clamshells are placed inside a 10.8-cup SnapWareairtight container (Model #109842). A 42.5 mm Whatman #1 filter disk(Cat. No. 1001-042) is placed on a watch glass. Compound A is dissolvedin acetone and added to the disks in a manner to produce a finalheadspace concentration of 50 mg/L. The acetone is permitted toevaporate for five minutes. The containers are then closed with thelids, and placed for three days at 21° C. After storage, fruits areevaluated for disease incidence (mm diameter of the rot) and pathogensporulation (mm diameter) on the surface of the fruits for an additionalthree days at 21° C., with results summarized in Table 19. Resultsdemonstrate 100% in vivo volatile antimicrobial control of P. expansummold of apple up to 3 days after treatment.

TABLE 19 Incidence and severity of Penicillium expansum on apples asdepicted by a brown rot and fungal spores on the surface of the fruitsCompound A Rot (mm) Sporulation (mm) Rate (mg/L) Day 0 Day 1 Day 2 Day 3Day 0 Day 1 Day 2 Day 3 50 0 0 0 0 0 0 0 0 Control 15.9 20.1 25.7 30 3.53.9 3.9 6.5

Example 21

In order to assess the in vivo activity of volatile antimicrobialCompound B in fruit, a volatile bioassay is developed using orange. Twooranges per replicate are placed inside a clamshell. Three fresh woundsper orange are inoculated with 30 μL of 1×10⁶ per mL Penicilliumdigitatum spore suspension. The clamshells are placed inside a 10.8-cupSnapWare airtight container (Model #109842). Compound B powder isintroduced to the containers by a sublimation device (copper tube heatedto 200° C. with fan flow at 0.5 L/min) to achieve a final headspaceconcentration of 0.4, 2, 10, or 50 mg/L. The containers are then closedwith the lids and placed for three days at 21° C. After storage, fruitsare evaluated for disease incidence (mm diameter of the rot) andpathogen sporulation (mm diameter) on the surface of the fruits for anadditional three days at 21° C., with results summarized in Table 20.Results demonstrate good in vivo volatile inhibition of P. digitatum inorange at rates of 0.4 mg/L and complete inhibition at 10 mg/L.

TABLE 20 Incidence and severity of Penicillium digitatum on oranges asdepicted by water soaked lesion and fungal spores on the surface of thefruits after a treatment with Compound B. Compound B Water soakedlesions (mm) Sporulation (mm) Rate (mg/L) Day 0 Day 1 Day 2 Day 3 Day 0Day 1 Day 2 Day 3 50 0 0 0 0 0 0 0 0 10 0 0 0 0.8 0 0 0 0 2 0.5 7.8 30.742.6 0 0.3 2.8 5.7 0.4 5.7 29.4 49.3 63.4 0.7 1.4 8.1 27.2 Control 12.335.5 61.1 83.2 0.3 2.7 8.5 44.5

Example 22

To assess the in vivo activity of volatile antimicrobial Compound A(FIG. 1) in fruit, a volatile bioassay is developed using apple, pear,orange, strawberry, grape and blueberry. Two apples, 2 oranges, 2 pears,8 strawberries, 16 grapes or 30 blueberries (per replicate, induplicate) are placed in a clamshell with the stem end facing up for allfruits except for strawberry (stem end facing downwards). A fresh woundis inoculated with 20 μL 1×10⁶ per mL Penicillium expansum sporesuspension (apple and pear), 20 μL 1×10⁶ per mL Penicillium digitatumspore suspension (orange), and 20 μL (strawberry and grape) or 10 μL(blueberry) of 1×10⁶ per mL Botrytis cinerea spore suspension. Theclamshells are placed inside a 117 L Rubbermaid storage box (Cat #2244)with lids closed. Compound A, dissolved in acetone, is pipetted onto acotton strip, where the acetone is allowed to evaporate for fiveminutes, and then introduced into the container by a sublimation device(copper tube heated to 200° C. with fan flow at 0.5 L/min) to achieve afinal headspace concentration of 10 mg/L. The containers are then heldfor three days at 21° C. After treatment, fruits are held for anadditional three days at 21° C., then evaluated for disease incidence(mm diameter of browning or water-soaked lesions) and pathogensporulation (mm diameter) for apple, pear and orange, as well asBotrytis cinerea disease incidence (%) and severity (0 to 4) forstrawberry, grape and blueberry, with results summarized in Table 21.Results demonstrate good in vivo antimicrobial control of at least threefungal pathogens on at least six different hosts when applied as avolatile fungicide.

TABLE 21 Effects of subliming Compound A as reflected by incidence andseverity of B. cinerea on strawberry, grape and blueberry, and severityon oranges, apples and pears as depicted by water soaked lesions,browning and sporulation after a three day treatment plus an additionalthree days at 21° C. Treatment Incidence (%) Severity (0-4) (10 mg/L)Strawberry Blueberry Grape Strawberry Blueberry Grape Compound A 18.8 526.7 0.09 0.03 0.1 Control 100 100 80 3.6 2.2 0.9 Water soaked lesionBrowning (mm) Sporulation (mm) Orange Apple Pear Apple Orange PearCompound A 3.04 5.4 2.7 0 8.55 0 Control 50.5 11.5 23.3 4.8 33.2 15.5

Example 23

To compare the ability of Compound A when actively volatilized bydifferent mechanisms, an in vivo assay using strawberry is performed.Eight strawberries are placed in a clamshell with stem end facingdownwards. A fresh wound is inoculated with 20 μL of 1×10⁵ per mLBotrytis cinerea spore suspension. The clamshell is placed in a 10.8-cupSnapWare airtight container (Model #109842) and closed with the lids.Compound A is dissolved in acetone and volatilized through a sealable ½inch side port by an ES-100-H SmartFog system (Reno, Nev.).Alternatively, Compound A, dissolved in acetone, is pipetted onto acotton strip, where the acetone is allowed to evaporate for fiveminutes, and then introduced into the container by a sublimation device(copper tube heated to 200° C. with fan flow at 0.5 L/min) to achieve afinal headspace concentration of 10 mg/L. The fruits are stored forthree days at 21° C. After the three days of treatment, fruit are storedfor an additional three days at 21° C., and then evaluated for incidence(%) and severity of disease (0 to 4). Results are summarized in Table 22and demonstrate good antimicrobial activity against Botrytis cinerea inthis in vivo analysis, indicating that Compound A is an effectivevolatile antimicrobial.

TABLE 22 Effects of different volatile application methods of Compound Aas reflected by incidence and severity of Botrytis cinerea on strawberryafter a three day treatment plus an additional three days at 21° C.Treatments Incidence (%) Severity (0 to 4) Fog, 10 mg/L Compound A 6.30.03 Fog, control 62.5 1.6 Sublimation, 10 mg/L Compound A 0.0 0.0Sublimation, control 100.0 3.7

Example 24

An in vivo assay is used to evaluate the ability of Compound A tovolatilize from different materials and control fungal pathogens. Eightstrawberries are placed in a clamshell with stem end facing downwards. Afresh wound is inoculated with 20 μL of 1×10⁶ per mL Botrytis cinereaspore suspension. The clamshells are then placed in a 10.8-cup SnapWareairtight container (Model #109842). Compound A is dissolved in acetoneand then evenly sprayed onto cellulose paper and Tyvek® fabric at a rateof 200 milligrams per square meter (mg/m²). The acetone is allowed toevaporate. Likewise Compound A is dissolved in propylene glycol andevenly sprayed onto cellulose paper and Tyvek® fabric. No evaporation isattempted in this case.

TABLE 23 Effects of different films and subsequent release of Compound Aon the incidence and severity of Botrytis cinerea on strawberries aftera three day treatment and additional two day storage at 21° C. Rate(mg/L) Type of Film Incidence (%) Severity (0-4) 0.4 Cellulose Paper37.5 0.8 2 Cellulose Paper 37.5 0.7 10 Cellulose Paper 12.5 0.2 0.4Tyvek ® 31.3 0.5 2 Tyvek ® 6.3 0.1 10 Tyvek ® 6.3 0.3 Control No film100 2.5

Pieces of material are cut to the appropriate dimensions to deliver afinal headspace concentration of 0.4, 2, or 10 mg/L. The containers areclose, and placed for three days at 21° C. After treatment, fruit arestored for an additional two days at 21° C. and then evaluated forincidence (%) and severity (0 to 4) of disease, with the resultssummarized in Table 23. Results demonstrate good in vivo antimicrobialactivity of Compound A against Botrytis cinerea, with a reduction inincidence and severity at all rates, in a dose dependent manner, andthat the volatile compound can be released from different materials.

Example 25

An in vivo assay is used to evaluate the ability of Compound A tovolatilize from different materials and control fungal pathogens. Eightstrawberries are placed in a clamshell with the stem end facingdownwards. A fresh wound is inoculated with 20 μL of 1×10⁶ per mLBotrytis cinerea spore suspension. The clamshells are then placed in a10.8-cup SnapWare airtight container (Model #109842). As a substrate forCompound A, either a 42.5 mm Whatman #1 filter disk (Cat. No. 1001-042)placed on a watch glass or 10 square centimeter (cm²) pieces ofcardboard typically used for packaging strawberries was used. Compound Ais dissolved in acetone and either pipetted on the disk or painted onthe cardboard at a rate to achieve a final headspace concentration of0.4, 2, or 10 mg/L. The acetone is permitted to evaporate for fiveminutes. The containers are closed, and placed for three days at 21° C.After treatment, fruit are stored for an additional two days at 21° C.and then evaluated for incidence (%) and severity (0 to 4) of disease,with the results summarized in Table 24. Results demonstrate good invivo antimicrobial activity of Compound A against Botrytis cinerea, witha reduction in incidence and severity at all rates, in a dose dependentmanner, and that the volatile compound can be released from differentmaterials.

TABLE 24 Effects of different films and subsequent release of Compound Aon the incidence and severity of Botrytis cinerea on strawberries aftera three day treatment and additional two day storage at 21° C. Rate(mg/L) Type of Material Incidence (%) Severity (0-4) 10 Cardboard 25 0.22 Cardboard 37.5 0.3 0.4 Cardboard 87.5 0.9 Control Cardboard 93.8 2.710 Filter Paper 18.8 0.3 2 Filter Paper 37.5 0.6 0.4 Filter Paper 56.32.5 Control Filter Paper 100 2.5

Example 26

An in vitro assay is used to evaluate the ability of Compound A (FIG. 1)to volatilize from different materials and control fungal growth.

TABLE 25 Effects of different materials on the volatile release ofCompound A and the subsequent in vitro inhibition (MIC) of Botrytiscinerea. Material MIC (mg/L) Polyethylene 0.28 PTFE-Coated Fiberglass0.56 Fiberglass 0.56 Cellulose 0.56 Silica 0.56 Aramid and Fiberglass0.56 Vinyl-Coated Polyester 0.56 Acrylic-Coated Fiberglass 0.56Silicone-Coated Fiberglass 0.56 PTFE 1.1 Cardboard 2.2 Aramid 2.2

PTFE-Coated (8577K81), Fiberglass (8816k1), Silica (8799K3), Aramid andFiberglass (8821K4), Vinyl-Coated Polyester (8843K31), Acrylic-CoatedFiberglass (8838K2), Silicone-Coated Fiberglass (87815K1), Aramid(McMaster-Carr, Santa Fe Springs, Calif.-1206T1), Polyethylene PCRsealing film, Cellulose (Whatman #1, Cat No. 1001-0155), and Cardboardare cut into disks of 15 mm diameter. 12-Well (6.5 mL volume per well)microtiter plates are used for the in vitro inhibition assay forvolatile antimicrobial compounds. A 3-mL volume of full-strength PotatoDextrose Agar (PDA) is added to each well. After cooling, 1 μL of 1×10⁵per mL Botrytis cinerea spore suspension is spot-pipetted to the centerof the agar. Plates are inoculated immediately prior to volatilefungicide treatment. The various materials are placed, in duplicate, onthe underside of a polyethylene PCR plate sealing film. Fordetermination of the minimum inhibitory concentration (MIC), compoundsare diluted in acetone, and the appropriate amount of compound is addedto the materials in a dose dependent manner to achieve a final headspaceconcentration of 35.7 to 0.03 mg/L. The acetone is permitted toevaporate for five minutes. The headspace around the Botrytis cinereainoculum is then sealed inside the well by the film with the adheringdisk of material containing the fungicide. Plates are inverted, placedover the treated disks and sealed to prevent any of the chemical fromflaking from the disk and falling onto the inoculated agar. After threedays of storage at 23° C., the cultures are evaluated for percent growthrelative to control based on measurement of fungal colony diameter.Experimental results are summarized in Table 25. The results indicatethat Compound A can volatilize from numerous materials to inhibit the invitro growth of Botrytis cinerea with similar levels of control.

Example 27

An in vivo assay is used to evaluate the ability of Compound A tocontrol fungal growth of seeds.

TABLE 26 Effect of a 10 mg/L headspace treatment of Compound A incontrolling Aspergillus brasiliensis growth on grains. Fungal growth onPDA (mm) Grains Compound A Control-Acetone Control-No Acetone Barley 012.8 21.7 Corn Dry 0 10.1 22.8 Millet 0 7.2 19.1 Rice 0 7.5 21.6 Rye 08.4 21 Wheat 0 8.1 22.4

Grains consisting of corn, wheat, rice, rye, millet and barley aresurface sterilized with 0.825% NaOCl for 1 minute and rinsed thrice withsterile distilled water. The grains are inoculated by soaking them in a1×10⁶ spores/mL suspension of Aspergillus brasiliensis for 1 minute. Theexcess inoculum is blotted out with a sterile paper towel before platingfive seeds in a Petri plate containing 25 mL of PDA. For determinationof efficacy, Compound A is diluted in acetone and added to 42.5 mmWhatman #1 filter disks (Cat. No. 1001-042) attached to the inner sideof the lid in a dose dependent manner to achieve a final headspaceconcentration of 0.4, 2, or 10 mg/L. The acetone is permitted toevaporate for five minutes before closing plate and sealing it withparafilm. The plates are incubated at 23° C. for three days. Afterstorage, the grains are evaluated for mycelial colony diameter (mm),with results summarized in Table 26. Results demonstrate 100% control ofAspergillus brasiliensis in this in vivo analysis.

Example 28

To evaluate a combination treatment of Compound A with1-methylcyclopropene (1-MCP), an in vivo experiment is performed onwhite roses.

TABLE 27 Botrytis cinerea incidence and severity based on infection onpetals and sepals of white roses after a 24 hours treatment with 1-MCPfollowed by a three day volatile treatment of Compound A at 21° C. andan additional five days at 21° C. Treatments Incidence (%) Severity*(0-4) Control 66.7 2.0 1-MCP 33.3 0.4 0.008 mg/L 20.0 0.2 0.04 mg/L 20.00.03 0.2 mg/L 0.0 0.0 0.008 mg/L + 1-MCP 6.7 0.9 0.04 mg/L + 1-MCP 0.00.0 0.2 mg/L + 1-MCP 0.0 0.0 *Severity Rating 0 = No disease 1 =Browning and small lesions on the sepals or petals 2 = Browning, petalscovered with fungal spores 3 = Browning, petals covered with fungalspores, some petal drop 4 = Browning, petals covered with fungal spores,some flowers aborted

Five white roses are placed in an 800 mL jar containing 200 mL of acommon commercial flower preservative. Three jars are then placed in a117 L Rubbermaid storage box (Cat #2244). Two small fans are placed inopposite ends of the container to assist with the distribution of thetwo volatiles. A 500 parts per billion (ppb) volume per volume (v/v)1-MCP treatment is applied (AgroFresh, Springhouse, Pa.) for 24 hours at21° C. After the 1-MCP treatment is completed, the containers arevented, and Compound A powder is applied in a dose dependent manner toachieve a final headspace concentration of 0.2, 0.04, or 0.008 mg/L witha sublimation device (copper tube heated to 200° C. with fan flow at 0.5L/min), with the end of the tube penetrating through a ½ inch side portin the container that is sealed immediately after the application. Theflowers are incubated for three days at 21° C. After treatment, theflowers are evaluated for an additional seven days at 21° C. for diseaseincidence and severity of the flower petals. Results listed in Table 27show good antimicrobial activity against Botrytis cinerea infection ofwhite roses, and that enhancing the rate of volatilization throughsublimation resulted in greater disease control. Also treatment with1-MCP shows reduced petal drop as reflected by severity scores.

Example 29

To evaluate a combination treatment of Compound A with1-methylcyclopropene (1-MCP), an in vivo experiment is performed onbroccoli.

TABLE 28 Effects of Compound A and 1-MCP in controlling Alternaria rotand yellowing of broccoli, respectively, five or three days treatment at10 or 21° C. with additional two days at 21° C. 21° C. 11 C. Treatments(mg/L) Severity Color Score* Severity Color Score* Control 1.5 2.39 0.181.55 1-MCP 0.61 1.79 0.18 1.50 0.4 mg/L 0.29 1.32 0.00 1.75 2 mg/L 0.071.89 0.00 2.11 0.4 mg/L + 1-MCP 0.21 0.93 0.00 1.39 2 mg/L + 1-MCP 0.071.93 0.00 2.23 Color Score Rating 0 = green, regular looking broccoli 1= Few light green spots 2 = Light green and yellow spots 3 = Lightgreen, yellow and some brown 4 = Mostly yellow and brown

Broccoli flowers are inoculated with 1×10⁶ spores/mL of Alternariaalternata and then placed in a 117 L Rubbermaid storage box (Cat #2244)with two small fans placed in opposite ends of the container. A 500 ppbv/v 1-MCP treatment is applied (AgroFresh, Springhouse, Pa.) for 24hours at 1° C. After completion of the 1-MCP treatment, broccoli floretsare removed and placed in a 10.8-cup SnapWare airtight container (Model#109842). Compound A powder is applied in a dose dependent manner toachieve a final headspace concentration of 2 or 0.4 mg/L with asublimation device (copper tube heated to 200° C. with fan flow at 0.5L/min), with the end of the tube penetrating through a ½ inch side portin the container that is sealed immediately after the application. Theflorets are incubated for five days at 10° C. or three days at 21° C.,then evaluated for an additional five days at 21° C. for diseaseincidence and severity. Results listed in Table 28 show goodantimicrobial activity against Alternaria alternata infection.

Example 30

To evaluate a combination treatment of Compound A with1-methylcyclopropene (1-MCP), an in vivo experiment is performed ontomato. Each tomato fruit is wounded three times and inoculated with1×10⁶ spores/mL of Alternaria alternata and then placed in a 117 LRubbermaid storage box (Cat #2244) with two small fans placed inopposite ends of the container. A 1000 ppb v/v 1-MCP treatment isapplied (AgroFresh, Springhouse, Pa.) for 24 hours at 21° C. Aftercompletion of the 1-MCP treatment, the tomatoes are removed and placedin a 10.8-cup SnapWare airtight container (Model #109842). Compound Apowder is applied in a dose dependent manner to achieve a finalheadspace concentration of 2 or 0.4 mg/L with a sublimation device(copper tube heated to 200° C. with fan flow at 0.5 L/min), with the endof the tube penetrating through a ½ inch side port in the container thatis sealed immediately after the application. The tomatoes are incubatedfor three days at 21° C., then evaluated for an additional three days at21° C. for disease incidence and severity. Results listed in Table 29show good activity against Alternaria alternata infection of tomato.

TABLE 29 Effects of Compound A and 1-MCP in controlling Alternaria roton tomatoes, three days treatment at 21° C. with additional three daysat 21° C. Compound A Diameter of the rot (mm) Control 14.8 1-MCP 13.60.4 mg/L 3.8 2 mg/L 0.0 0.4 mg/L + 1-MCP 3.8 2 mg/L + 1-MCP 0.0

Example 31

To assess the in vivo activity of volatile antimicrobial Compounds A andB (FIG. 1) in fruit, a volatile bioassay is developed using apple, pear,orange, strawberry, grape and blueberry.

TABLE 30 Effects of subliming Compounds A and B as reflected byincidence and severity of B. cinerea on strawberry, grape and blueberry,and severity on oranges, apples and pears as depicted by water soakedlesions, browning and sporulation after a three day treatment plus anadditional three days at 21° C. Treatments Incidence (%) Severity (0-4)(1 mg/L) Strawberry Blueberry Grape Strawberry Blueberry Grape CompoundA 0.0 0.0 0.0 0.0 0.0 0.0 Compound B 0.0 0.0 0.0 0.0 0.0 0.0 Control100.0 100.0 100.0 3.9 2.5 1.9 Water soaked lesion Browning (mm)Sporulation (mm) Orange Apple Pear Apple Orange Pear Compound A 0.0 0.84.7 0.0 0.0 0.0 Compound B 0.0 2.3 1.1 0.2 0.0 0.0 Control 73.2 21.729.7 46.0 5.2 18.5

Two apples, 2 oranges, 2 pears, 8 strawberries, 16 grapes or 30blueberries (per replicate, in duplicate) are placed in a clamshell withthe stem end facing up for all fruits except for strawberry (stem endfacing downwards). A fresh wound is inoculated with 20 μL 1×10⁶ per mLPenicillium expansum spore suspension (apple and pear), 20 μL 1×10⁶ permL Penicillium digitatum spore suspension (orange), and 20 μL(strawberry and grape) or 10 μL (blueberry) of 1×10⁶ per mL Botrytiscinerea spore suspension. The clamshells are placed inside a 117 LRubbermaid storage box (Cat #2244) with lids closed. Compound A and Bpowders are introduced to the containers by a sublimation device (coppertube heated to 200° C. with fan flow at 0.5 L/min) to achieve a finalheadspace concentration of 1 mg/L. The containers are then held forthree days at 21° C. After treatment, fruits are held for an additionalthree days at 21° C., then evaluated for disease incidence (mm diameterof browning or water-soaked lesions) and pathogen sporulation (mmdiameter) for apple, pear and orange, as well as Botrytis cinereadisease incidence (%) and severity (0 to 4) for strawberry, grape andblueberry, with results summarized in Table 30. Results demonstrate 100%in vivo antimicrobial control of B. cinerea and P. digitatum by bothCompounds A and B on different hosts when applied as a volatilefungicide.

Example 32

In order to assess the activity of Compound A as a contact fungicide, anin vitro assay is developed. A 6 cm-diameter Petri plate is used.Compound A is amended into full-strength Potato Dextrose Agar (PDA) toachieve a final solution concentration of 10, 2, 0.4, or 0.08 mg/L, and15-mL volume of solution is added to each plate. After cooling, 1 μL of1×10⁵ per mL Penicillium expansum or Penicillium digitatum sporesuspension is spot-pipetted into the center of the agar.

Plates are sealed with a parafilm and placed in an incubator held at 23°C. After three days of storage, the cultures are evaluated for percentgrowth relative to control based on measurement of fungal colonydiameter. Experimental results are summarized in Table 31. The resultsindicate that Compound A has activity as a contact fungicide in this invitro assay against plant fungal pathogens.

TABLE 31 The in vitro MIC for Compound A as a contact fungicide formycelial growth inhibition of Penicillium expansum and Penicilliumdigitatum. Pathogen Incidence (%) Rate (mg/L) P. expansum P. digitatum10 0.0 0.0 2 0.0 0.0 0.4 33.0 12.5 0.08 93.1 42.0

Example 33

In order to assess the activity of Compound A (FIG. 1) as a contactdrench fungicide, an in vivo assay is developed. Two apples or 2 oranges(per replicate, in duplicate) are placed in a clamshell, and three freshwounds near the equatorial region of each fruit are made. Compound A isdissolved in water to achieve a final treatment solution concentrationof 250, 50, or 10 mg/L. The fruit is dipped in Compound A solution for 1minute and allowed to dry for 1 hour. Fruit wounds are then inoculatedwith 30 μL of 1×10⁶ per mL Penicillium expansum spore suspension (apple)or Penicillium digitatum spore suspension (orange). Clamshells are thenplaced in a 10.8-cup SnapWare airtight container (Model #109842) andincubated for 3 days at 21° C. After treatment, the fruit is held for anadditional 3 days at 21° C. and then evaluated for disease incidence (mmdiameter of browning or water-soaked lesions) and pathogen sporulation(mm diameter), with results summarized in Table 32. Results demonstrategood in vivo antimicrobial control of 2 fungal pathogens on 2 differenthosts when applied as a contact fungicide.

TABLE 32 In vivo MIC for Compound A as a contact fungicide for controlof Penicillium digitatum and Penicillium expansum on oranges and applesrespectively. Oranges Apples Compound A Water soaked SporulationBrowning Sporulation (mg/L) (mm) (mm) (mm) (mm) Control 42.7 31.0 9.73.5 10 27.5 16.6 8.5 2.1 50 18.8 12.0 3.7 1.6 250  1.7 0.4 0.8 0.5

Example 34

In order to assess the activity of Compound A as a volatile fungicide,an in vitro assay is developed to evaluate spore germination. Two mL ofwater agar is poured in 3.5 cm Petri plates. Compound A is dissolved inacetone to achieve a final treatment solution concentration of 0.14,0.07, or 0.035 mg/L. Plates are inoculated with 1 μL 1×10⁶ per mLBotrytis cinerea and Penicillium expansum spore suspension. Plates arethen incubated for either one day at 0° C., five day at 0° C., or fiveday at 0° C. plus an additional one or two days at 21° C. At each timepoint, plates are removed and 100 spores are counted for percentgermination, where germination is defined as a germ tube that hasextended a distance greater than the length of the spore. Results aresummarized in Table 33. At all three treatment concentrations andtemperature regimes, Compound A completely inhibits the germination ofthe fungal pathogens spores tested.

TABLE 33 Percent germination of Botrytis cinerea and Penicilliumexpansum spores in response to a volatile treatment with Compound Aunder 4 different temperature regimes Germination Inhibition (%) 1 5 5day, 0° C. 5 day, 0° C. Compound A day, day, 1 day, 2 day, PathogensRate (mg/L) 0° C. 0° C. 21° C. 21° C. B. cinerea 0.14 0.0 0.0 0.0 0.00.07 0.0 0.0 0.0 0.0 0.035 0.0 0.0 0.0 0.0 Control 44.8 98.7 92.2 98.4Acetone 48.9 98.9 93.9 95.8 P. expansum 0.14 0.0 0.0 0.0 0.0 0.07 0.00.0 0.0 0.0 0.035 0.0 0.0 0.0 0.0 Control 0.0 1.1 12.6 30.8 Acetone 0.00.0 6.4 21.8

Example 35

In order to assess the activity of Compound A as a volatile fungicide,an in vitro assay is developed to evaluate spore germination. 3.5-cmPetri plates are filled with 2 mL of water agar. After cooling, 1 μL of1×10⁵ per mL Botrytis cinerea spore suspension is spot-pipetted into thecenter of the plate. Plates are inoculated immediately prior to volatilefungicide treatment. A Whatman #1 filter disk (Cat. No. 1001-0155) isplaced, in duplicate, on the underside of a plate lid. For determinationof the minimum inhibitory concentration (MIC), compounds are diluted inacetone, and the appropriate amount of compound is added to the disks ina dose dependent manner to achieve a final headspace concentration of142.9 to 0.07 mg/L. The acetone is permitted to evaporate for fiveminutes, and then the lids are placed on the plates and sealed withparafilm. After 24 hours of storage at 23° C., 100 spores are countedfor percent germination, where germination is defined as a germ tubethat has extended a distance greater than the length of the spore. Aftercounting, treatment is removed, and the plates are resealed. After anadditional 24 hours, 100 spores are again counted. Plugs are thentransferred to a clean plate containing full-strength PDA and allowed toincubate at 23° C. for an additional three days. After incubation,mycelial growth (mm diameter) is determined and summarized in Table 34.After 24 hours, 100% of the control spores have germinated while allrates of Compound A resulted in 100% inhibition of germination in thisvolatile in vitro assay. These results show that Compound A delivers afungicidal effect, as opposed to a fungistatic effect, such that treatedspores fail to germinate and grow as mycelia even after the compound hasbeen removed.

TABLE 34 Spore germination and subsequent mycelial growth after transferto fresh media of Botrytis cinerea in response to a volatile treatmentof Compound A. Spore germination Mycelial growth Compound A (%) aftertransfer (%) Rate (mg/L) 24 h^(a) 24 h + 24 h^(b) 3 d^(c) Control 100.0100.0 100.0 Acetone 98.4 92.7 100.0 142.9  0.0 0.0 0.0 71.4  0.0 0.0 0.035.7  0.0 0.0 0.0 17.9  0.0 0.0 0.0 8.9 0.0 0.0 0.0 4.5 0.0 0.0 10.1 2.20.0 0.0 16.9 1.1 0.0 0.0 32.6  0.56 0.0 0.0 43.3  0.28 0.0 0.0 51.3 0.14 0.0 0.0 53.8  0.07 0.0 10.0 60.3 ^(a)Spore germination determinedafter 24 hours treatment ^(b)Spore germination determined afteradditional 24 hours after treatment removal ^(c)Percent mycelial growth3 days after transfer of inoculum to clean PDA plates

We claim:
 1. A method of using a volatile antimicrobial compound againstpathogens affecting meats, plants, or plant parts, comprising providingin gaseous form a volatile antimicrobial compound of formula (IV):

contacting a meat, plant, or plant part with an effective amount of thevolatile antimicrobial compound in gaseous form; and contacting themeat, plant, or plant part with an effective amount of a cyclopropenecompound in gaseous form; wherein A and D together with the carbon atomsto which they are attached form a 5-, 6-, or 7-membered fused ring whichmay be substituted by C₁-C₆-alkyl, C₁-C₆-alkoxy, hydroxy, halogen,nitro, nitrile, amino, amino substituted by one or more C₁-C₆-alkylgroups, carboxy, acyl, aryloxy, carbonamido, carbonamido substituted byC₁-C₆-alkyl, sulfonamido or trifluoromethyl or the fused ring may linktwo oxaborole rings; X is a group —CR⁷R⁸ wherein R⁷ and R⁸ are eachindependently hydrogen, C₁-C₆-alkyl, nitrile, nitro, aryl, arylalkyl orR⁷ and R⁸ together with the carbon atom to which they are attached forman alicyclic ring; and R⁶ is hydrogen, C₁-C₁₈-alkyl, C₁-C₁₈-alkylsubstituted by C₁-C₆-alkoxy, C₁-C₆-alkylthio, hydroxy, amino, aminosubstituted by C₁-C₁₈-alkyl, carboxy, aryl, aryloxy, carbonamido,carbonamido substituted by C₁-C₆-alkyl, aryl or arylalkyl, arylalkyl,aryl, heteroaryl, cycloalkyl, C₁-C₁₈-alkyleneamino, C₁-C₁₈-alkyleneaminosubstituted by phenyl, C₁-C₆-alkoxy or C₁-C₆-alkylthio, carbonylalkyleneamino or a radical of formula (V):

wherein A, D and X are as defined herein before except forboronophthalide; and agriculturally acceptable salts thereof.
 2. Themethod of claim 1, wherein the pathogen is at least one member selectedfrom the group consisting of Acremonium spp., Albugo spp., Alternariaspp., Ascochyta spp., Aspergillus spp., Bacillus spp., Botryodiplodiaspp., Botryospheria spp., Botrytis spp., Byssochlamys spp.,Campylobacter spp., Candida spp., Cephalosporium spp., Ceratocystisspp., Cercospora spp., Chalara spp., Cladosporium spp., Clavibacterspp., Clostridium spp., Colletotrichum spp., Cryptosporidium spp.,Cryptosporiopsis spp., Cylindrocarpon spp., Debaryomyces spp., Diaporthespp., Didymella spp., Diplodia spp., Dothiorella spp., Elsinoe spp.,Erwinia spp., Escherichia spp., Fusarium spp., Geotrichum spp., Giardiaspp., Gloeosporium spp., Glomerella spp., Helminthosporium spp., Khuskiaspp., Lactobacillus spp., Lasiodiplodia spp., Leuconostoc spp., Listeriaspp., Macrophoma spp., Macrophomina spp., Microdochium spp., Moniliniaspp., Monilochaethes spp., Mucor spp., Mycocentrospora spp.,Mycosphaerella spp., Nectria spp., Neofabraea spp., Nigrospora spp.,Pantoea spp., Pectobacterium spp., Penicillium spp., Peronophythoraspp., Peronospora spp., Pestalotiopsis spp., Pezicula spp.,Phacidiopycnis spp., Phoma spp., Phomopsis spp., Phyllosticta spp.,Phytophthora spp., Polyscytalum spp., Pseudocercospora spp., Pseudomonasspp., Pyricularia spp., Pythium spp., Ralstonia spp., Rhizoctonia spp.,Rhizopus spp., Salmonella spp., Sclerotium spp., Sclerotinia spp.,Septoria spp., Shigella spp., Sphaceloma spp., Sphaeropsis spp.,Staphylococcus spp., Stemphyllium spp., Stilbella spp., Thielaviopsisspp., Thyronectria spp., Trachysphaera spp., Uromyces spp., Ustilagospp., Venturia spp., Verticillium spp., Vibrio spp., Xanthomonas spp.,and Yersinia spp.
 3. The method of claim 1, wherein the method comprisesa treatment selected from the group consisting of treatment during fieldpacking, treatment in clamshells, treatment during palletization orafter palletization, treatment in open pallets or in wrapped pallets,treatment in tents, treatments inside boxes with or without liners, insea container, truck or other container types used duringtransportation, and treatment during storage.
 4. The method of claim 1,wherein the plants or plant parts are selected from the group consistingof asparagus, sugar beet, barley, broccoli, cabbage, carrot, cassava,cauliflower, celery, cucumber, eggplant, garlic, grapevine, lettuce,spinach, leek, mushroom, onion, peas, pepper, bell pepper, potato,pumpkin, rye, sweet potato, squash, tobacco, tomato, snap bean, sorghum,sugarcane, corn, wheat, cotton, rice, soybean, canola, fruit,vegetables, nursery, turf, flowers, carnation, geranium, lily, orchid,rose, sunflower, ornamental flowers and ornamental crops.
 5. The methodof claim 4, wherein the fruit is selected from the group consisting ofapple, avocado, banana, strawberry, blueberry, raspberry, blackberry,cherry, oranges, lemon, lime, grapefruit, fig, grapes, guava, kiwifruit,mango, nectarine, cantaloupe, muskmelon, watermelon, papaya, peach,pear, persimmon, pineapple, and pomegranate.
 6. The method of claim 4,wherein the meat is selected from the group consisting of beef, bison,chicken, deer, goat, turkey, pork, sheep, fish, shellfish, mollusks anddry-cured meat products.
 7. The method of claim 1, wherein thecontacting comprises applying the volatile antimicrobial compound by agas treatment selected from the group consisting of release from asachet, release from a synthetic or natural film, release from liner orother packaging materials, release from powder, release from agas-releasing generator, release using a compressed or non-compressedgas cylinder, release from a droplet or droplets placed inside a box,release from a mist or fog applied into a container and combinationsthereof.
 8. The method of claim 1, wherein the volatile antimicrobialcompound has a structure of


9. A method of using a volatile antimicrobial compound against pathogensaffecting meats, plants, or plant parts, comprising placing a meat,plant or plant part in a container; introducing into the container andin contact with the meat, plant or plant part an effective amount of agaseous form of a volatile antimicrobial compound of formula (IV):

wherein A and D together with the carbon atoms to which they areattached form a 5-, 6-, or 7-membered fused ring which may besubstituted by C₁-C₆-alkyl, C₁-C₆-alkoxy, hydroxy, halogen, nitro,nitrile, amino, amino substituted by one or more C₁-C₆-alkyl groups,carboxy, acyl, aryloxy, carbonamido, carbonamido substituted byC₁-C₆-alkyl, sulfonamido or trifluoromethyl or the fused ring may linktwo oxaborole rings; X is a group —CR⁷R⁸ wherein R⁷ and R⁸ are eachindependently hydrogen, C₁-C₆-alkyl, nitrile, nitro, aryl, arylalkyl orR⁷ and R⁸ together with the carbon atom to which they are attached forman alicyclic ring; and R⁶ is hydrogen, C₁-C₁₈-alkyl, C₁-C₁₈-alkylsubstituted by C₁-C₆-alkoxy, C₁-C₆-alkylthio, hydroxy, amino, aminosubstituted by C₁-C₁₈-alkyl, carboxy, aryl, aryloxy, carbonamido,carbonamido substituted by C₁-C₆-alkyl, aryl or arylalkyl, arylalkyl,aryl, heteroaryl, cycloalkyl, C₁-C₁₈-alkyleneamino, C₁-C₁₈-alkyleneaminosubstituted by phenyl, C₁-C₆-alkoxy or C₁-C₆-alkylthio, carbonylalkyleneamino or a radical of formula (V):

wherein A, D and X are as defined herein before except forboronophthalide; and agriculturally acceptable salts thereof; andintroducing into the container and in contact with the meat, plant orplant part an effective amount of 1-methylcyclopropene (1-MCP).
 10. Themethod of claim 9, wherein the method comprises a treatment selectedfrom the group consisting of treatment during field packing, treatmentin clamshells, treatment during palletization or after palletization,treatment in open pallets or in wrapped pallets, treatment in tents,treatments inside boxes with or without liners, in sea container, truckor other container types used during transportation, and treatmentduring storage.
 11. The method of claim 9, wherein the plants or plantparts are selected from the group consisting of asparagus, sugar beet,barley, broccoli, cabbage, carrot, cassava, cauliflower, celery,cucumber, eggplant, garlic, grapevine, lettuce, spinach, leek, mushroom,onion, peas, pepper, bell pepper, potato, pumpkin, rye, sweet potato,squash, tobacco, tomato, snap bean, sorghum, sugarcane, corn, wheat,cotton, rice, soybean, canola, fruit, vegetables, nursery, turf,flowers, carnation, geranium, lily, orchid, rose, sunflower, ornamentalflowers and ornamental crops.
 12. The method of claim 11, wherein thefruit is selected from the group consisting of apple, avocado, banana,strawberry, blueberry, raspberry, blackberry, cherry, oranges, lemon,lime, grapefruit, fig, grapes, guava, kiwifruit, mango, nectarine,cantaloupe, muskmelon, watermelon, papaya, peach, pear, persimmon,pineapple, and pomegranate.
 13. The method of claim 9, wherein the meatis selected from the group consisting of beef, bison, chicken, deer,goat, turkey, pork, sheep, fish, shellfish, mollusks and dry-cured meatproducts.
 14. The method of claim 9, wherein the contacting comprisesapplying the volatile antimicrobial compound by a gas treatment selectedfrom the group consisting of release from a sachet, release from asynthetic or natural film, release from liner or other packagingmaterials, release from powder, release from a gas-releasing generator,release using a compressed or non-compressed gas cylinder, release froma droplet or droplets placed inside a box, release from a mist or fogapplied into a container and combinations thereof.
 15. The method ofclaim 9, wherein the volatile antimicrobial compound has a structure of


16. A method of treating plants, or plant parts, comprising contactingthe plants or plant parts with an atmosphere comprising an effectiveamount of a volatile antimicrobial compound in gaseous form, wherein thevolatile antimicrobial compound is a compound of formula (IV):

wherein A and D together with the carbon atoms to which they areattached form a 5-, 6-, or 7-membered fused ring which may besubstituted by C₁-C₆-alkyl, C₁-C₆-alkoxy, hydroxy, halogen, nitro,nitrile, amino, amino substituted by one or more C₁-C₆-alkyl groups,carboxy, acyl, aryloxy, carbonamido, carbonamido substituted byC₁-C₆-alkyl, sulfonamido or trifluoromethyl or the fused ring may linktwo oxaborole rings; X is a group —CR⁷R⁸ wherein R⁷ and R⁸ are eachindependently hydrogen, C₁-C₆-alkyl, nitrile, nitro, aryl, arylalkyl orR⁷ and R⁸ together with the carbon atom to which they are attached forman alicyclic ring; and R⁶ is hydrogen, C₁-C₁₈-alkyl, C₁-C₁₈-alkylsubstituted by C₁-C₆-alkoxy, C₁-C₆-alkylthio, hydroxy, amino, aminosubstituted by C₁-C₁₈-alkyl, carboxy, aryl, aryloxy, carbonamido,carbonamido substituted by C₁-C₆-alkyl, aryl or arylalkyl, arylalkyl,aryl, heteroaryl, cycloalkyl, C₁-C₁₈-alkyleneamino, C₁-C₁₈-alkyleneaminosubstituted by phenyl, C₁-C₆-alkoxy or C₁-C₆-alkylthio, carbonylalkyleneamino or a radical of formula (V):

wherein A, D and X are as defined herein before except forboronophthalide; and agriculturally acceptable salts thereof; andwherein the atmosphere further comprises an effective amount of1-methylcyclopropene (1-MCP).
 17. The method of claim 16, wherein thevolatile antimicrobial compound has a structure of


18. The method of treatment of claim 16, wherein the method comprises atreatment selected from the group consisting of treatment during fieldpacking, treatment in clamshells, treatment during palletization orafter palletization, treatment in open pallets or in wrapped pallets,treatment in tents, treatments inside boxes with or without liners, insea container, truck or other container types used duringtransportation, and treatment during storage.